Nucleic acid comprising or coding for a histone stem-loop and a poly(a) sequence or a polyadenylation signal for increasing the expression of an encoded allergenic antigen or an autoimmune self-antigen

ABSTRACT

The present invention relates to a nucleic acid sequence, comprising or coding for a coding region, encoding at least one peptide or protein comprising an allergenic antigen or an autoimmune self-antigen or a fragment, variant or derivative thereof, at least one histone stem-loop and a poly(A) sequence or a polyadenylation signal. Furthermore the present invention provides the use of the nucleic acid for increasing the expression of said encoded peptide or protein. It also discloses its use for the preparation of a pharmaceutical composition, especially a vaccine, e.g. for use in the treatment of allergies or autoimmune diseases. The present invention further describes a method for increasing the expression of a peptide or protein comprising an allergenic antigen or an autoimmune self-antigen or a fragment, variant or derivative thereof, using the nucleic acid comprising or coding for a histone stem-loop and a poly(A) sequence or a polyadenylation signal.

The present invention relates to a nucleic acid sequence, comprising orcoding for a coding region, encoding at least one peptide or proteincomprising an allergenic antigen or an autoimmune self-antigen or afragment, variant or derivative thereof, at least one histone stem-loopand a poly(A) sequence or a polyadenylation signal. Furthermore, thepresent invention provides the use of the nucleic acid for increasingthe expression of said encoded peptide or protein. It also discloses itsuse for the preparation of a pharmaceutical composition, especially avaccine, e.g. for use in the treatment of allergies or autoimmunediseases. The present invention further describes a method forincreasing the expression of a peptide or protein comprising anallergenic antigen or an autoimmune self-antigen or a fragment, variantor derivative thereof, using the nucleic acid comprising or coding for ahistone stem-loop and a poly(A) sequence or a polyadenylation signal.

An allergy is a hypersensitivity disorder of the immune system. Allergicreactions occur when a person's immune system reacts to normallyharmless substances in the environment. A substance that causes areaction is called an allergen or an allergenic antigen. Allergy is oneof four forms of hypersensitivity and is formally called type I (orimmediate) hypersensitivity. Allergic reactions are distinctive becauseof excessive activation of certain white blood cells called mast cellsand basophils by Immunoglobulin E (IgE). This reaction results in aninflammatory response which can range from uncomfortable to dangerous.Mild allergies like hay fever are very common in the human populationand cause symptoms such as red eyes, itchiness, and runny nose, eczema,hives, hay fever, or an asthma attack. Allergies can play a major rolein conditions such as asthma. In some people, severe allergies toenvironmental or dietary allergens or to medication may result inlife-threatening reactions called anaphylaxis. Food allergies andreactions to the venom of stinging insects such as wasps and bees areoften associated with these severe reactions. Treatments for allergiesinclude avoiding known allergens, use of medications such asanti-histamines that specifically prevent allergic reactions, steroidsthat modify the immune system in general, and medications such asdecongestants that reduce the symptoms. Newer approaches useimmunotherapy to desensitize the body's immune response.

Desensitization or hyposensitization is a treatment in which the patientis gradually vaccinated with progressively larger doses of the allergenin question. This can either reduce the severity or eliminatehypersensitivity altogether. It relies on the progressive skewing of IgGantibody production, to block excessive IgE production seen in atopys.In a sense, the person builds up immunity to increasing amounts of theallergen in question. Studies have demonstrated the long-term efficacyand the preventive effect of immunotherapy in reducing the developmentof new allergy.

But desensitization with protein allergens is associated with severalproblems, like e.g. crosslinking of pre-existing immunoglobulin E onmast cells/basophils or induction of de novo synthesis of immunoglobulinE by the protein immunization itself. Genetic immunization offersinnovative solutions to these major problems associated with proteinimmunization. It easily enables the routine production of hypoallergenicvaccines, which do not translate native allergens, thus avoidingpotential anaphylactic side effects. Genetic vaccines can also beapplied as mixtures of single vaccines, making them interestingcandidates for treatment based on component-resolved diagnosis, followedby an individualized therapy with the relevant allergens.

Autoimmune diseases typically arise from an overactive immune responseof the body against self-antigens (autoantigens), which are present andproduced in the body, i.e., the body attacks its own cells, proteins, orother components of the body. Autoimmune diseases as known today can beclassified as hypersensibility reactions according to known subtypes II,III or IV. Hypersensibility reactions of subtype I comprise allergiesand asthma. These reactions are directed against exogenous antigens andare therefore not regarded as autoimmune reactions.

In summary, autoimmunity may be recognized as the failure of an organismto discriminate between its own constituent parts (down to thesub-molecular levels) as self-antigens and (exogeneous)non-self-antigens, which typically induce an immune response in thebody. Furthermore, the components of the immune systems do not appear tobe capable to entirely delete the specific self-antigen or fragmentsthereof, thus resulting in (chronic) inflammatory autoimmune diseases.

Treatment of autoimmune diseases today is mainly restricted toalleviation of the symptoms rather than abolishment of the causes ofautoimmune diseases. Present approaches include, e.g., suppression ofthe body's immune response in order to alleviate the (inflammatory)attack on the own tissue and damage of tissue due to (inflammatory)attacks. Medicaments suitable for such a treatment are typicallyselected from immunosuppressive medicaments. However, an overallsuppression of immune system may lead to severe problems: Whenabolishing or diminishing the negative activities of the immune systemthe necessary protective activity of the body's immune system istypically impaired. If an immunosuppressive medicament is capable toefficiently protect attacked organs, the whole organism's capability tocombat infections is impaired. Up to now, there are only singleattempts, which try to circumvent these general problems.

Therefore, supplementary strategies have been investigated in recentyears in addition to such “conventional treatments” to avoid or at leastreduce the impact on the immune system by such treatments. One suchsupplementary treatment in particular includes gene therapeuticapproaches or genetic vaccination, which already have been found to behighly promising for treatment or for supporting such conventionaltherapies.

Gene therapy and genetic vaccination are methods of molecular medicinewhich already have been proven in the therapy and prevention of diseasesand generally exhibit a considerable effect on daily medical practice,in particular on the treatment of diseases as mentioned above. Bothmethods, gene therapy and genetic vaccination, are based on theintroduction of nucleic acids into the patient's cells or tissue andsubsequent processing of the information coded for by the nucleic acidthat has been introduced into the cells or tissue, that is to say the(protein) expression of the desired polypeptides.

In gene therapy approaches, typically DNA is used even though RNA isalso known in recent developments. Importantly, in all these genetherapy approaches mRNA functions as messenger for the sequenceinformation of the encoded protein, irrespectively if DNA, viral RNA ormRNA is used.

In general RNA is considered an unstable molecule: RNases are ubiquitousand notoriously difficult to inactivate. Furthermore, RNA is alsochemically more labile than DNA. Thus, it is perhaps surprising that the“default state” of an mRNA in a eukaryotic cell is characterized by arelative stability and specific signals are required to accelerate thedecay of individual mRNAs. The main reason for this finding appears tobe that mRNA decay within cells is catalyzed almost exclusively byexonucleases. However, the ends of eukaryotic mRNAs are protectedagainst these enzymes by specific terminal structures and theirassociated proteins: a m7GpppN CAP at the 5′ end and typically a poly(A)sequence at the 3′ end. Removal of these two terminal modifications isthus considered rate limiting for mRNA decay. Although a stabilizingelement has been characterized in the 3′ UTR of the alpha-globin mRNA,RNA sequences affecting turnover of eukaryotic mRNAs typically act as apromoter of decay usually by accelerating deadenylation (reviewed inMeyer, S., C. Temme, et al. (2004), Crit Rev Biochem Mol Biol 39(4):197-216).

As mentioned above, the 5′ ends of eukaryotic mRNAs are typicallymodified posttranscriptionally to carry a methylated CAP structure, e.g.m7GpppN. Aside from roles in RNA splicing, stabilization, and transport,the CAP structure significantly enhances the recruitment of the 40Sribosomal subunit to the 5′ end of the mRNA during translationinitiation. The latter function requires recognition of the CAPstructure by the eukaryotic initiation factor complex eIF4F. The poly(A)sequence additionally stimulates translation via increased 40S subunitrecruitment to mRNAs, an effect that requires the intervention ofpoly(A) binding protein (PABP). PABP, in turn, was recently demonstratedto interact physically with eIF4G, which is part of the CAP-bound eIF4Fcomplex. Thus, a closed loop model of translation initiation on capped,polyadenylated mRNAs was postulated (Michel, Y. M., D. Poncet, et al.(2000), J Biol Chem 275(41): 32268-76).

Nearly all eukaryotic mRNAs end with such a poly(A) sequence that isadded to their 3′ end by the ubiquitous cleavage/polyadenylationmachinery. The presence of a poly(A) sequence at the 3′ end is one ofthe most recognizable features of eukaryotic mRNAs. After cleavage, mostpre-mRNAs, with the exception of replication-dependent histonetranscripts, acquire a polyadenylated tail. In this context, 3′ endprocessing is a nuclear co-transcriptional process that promotestransport of mRNAs from the nucleus to the cytoplasm and affects thestability and the translation of mRNAs. Formation of this 3′ end occursin a two step reaction directed by the cleavage/polyadenylationmachinery and depends on the presence of two sequence elements in mRNAprecursors (pre-mRNAs); a highly conserved hexanucleotide AAUAAA(polyadenylation signal) and a downstream G/U-rich sequence. In a firststep, pre-mRNAs are cleaved between these two elements. In a second steptightly coupled to the first step the newly formed 3′ end is extended byaddition of a poly(A) sequence consisting of 200-250 adenylates whichaffects subsequently all aspects of mRNA metabolism, including mRNAexport, stability and translation (Dominski, Z. and W. F. Marzluff(2007), Gene 396(2): 373-90).

The only known exception to this rule are the replication-dependenthistone mRNAs which end with a histone stem-loop instead of a poly(A)sequence. Exemplary histone stem-loop sequences are described in Lopezet al. (Dávila López, M., & Samuelsson, T. (2008), RNA (New York, N.Y.),14(1), 1-10. doi:10.1261/rna.782308).

The stem-loops in histone pre-mRNAs are typically followed by apurine-rich sequence known as the histone downstream element (HDE).These pre-mRNAs are processed in the nucleus by a single endonucleolyticcleavage approximately 5 nucleotides downstream of the stem-loop,catalyzed by the L17 snRNP through base pairing of the U7 snRNA with theHDE. The 3′-UTR sequence comprising the histone stem-loop structure andthe histone downstream element (HDE) (binding site of the U7 snRNP) wereusually termed as histone 3′-processing signal (see e.g. Chodchoy, N.,N. B. Pandey, et al. (1991). Mol Cell Biol 11(1): 497-509).

Due to the requirement to package newly synthesized DNA into chromatin,histone synthesis is regulated in concert with the cell cycle. Increasedsynthesis of histone proteins during S phase is achieved bytranscriptional activation of histone genes as well asposttranscriptional regulation of histone mRNA levels. It could be shownthat the histone stem-loop is essential for all posttranscriptionalsteps of histone expression regulation. It is necessary for efficientprocessing, export of the mRNA into the cytoplasm, loading ontopolyribosomes, and regulation of mRNA stability.

In the above context, a 32 kDa protein was identified, which isassociated with the histone stem-loop at the 3′-end of the histonemessages in both the nucleus and the cytoplasm. The expression level ofthis stem-loop binding protein (SLBP) is cell-cycle regulated and ishighest during S-phase when histone mRNA levels are increased. SLBP isnecessary for efficient 3′-end processing of histone pre-mRNA by the U7snRNP. After completion of processing, SLBP remains associated with thestem-loop at the end of mature histone mRNAs and stimulates theirtranslation into histone proteins in the cytoplasm. (Dominski, Z. and W.F. Marzluff (2007), Gene 396(2): 373-90). Interestingly, the RNA bindingdomain of SLBP is conserved throughout metazoa and protozoa (DávilaLópez, M., & Samuelsson, T. (2008), RNA (New York, N.Y.), 14(1), 1-10.doi:10.1261/rna.782308) and it could be shown that its binding to thehistone stem-loop sequence is dependent on the stem-loop structure andthat the minimum binding site contains at least 3 nucleotides 5′ and 2nucleotides 3′ of the stem-loop (Pandey, N. B., et al. (1994), Molecularand Cellular Biology, 14(3), 1709-1720 and Williams, A. S., & Marzluff,W. F., (1995), Nucleic Acids Research, 23(4), 654-662).

Even though histone genes are generally classified as either“replication-dependent”, giving rise to mRNA ending in a histonestem-loop, or “replacement-type”, giving rise to mRNA bearing apoly(A)-tail instead, naturally occurring mRNAs containing both ahistone stem-loop and poly(A) or oligo(A) 3′ thereof have beenidentified in some very rare cases. Sanchez et al. examined the effectof naturally occurring oligo(A) tails appended 3′ of the histonestem-loop of histone mRNA during Xenopus oogenesis using Luciferase as areporter protein and found that the oligo(A) tail is an active part ofthe translation repression mechanism that silences histone mRNA duringoogenesis and its removal is part of the mechanism that activatestranslation of histone mRNAs (Sanchez, R. and W. F. Marzluff (2004), MolCell Biol 24(6): 2513-25).

Furthermore, the requirements for regulation of replication dependenthistones at the level of pre-mRNA processing and mRNA stability havebeen investigated using artificial constructs coding for the markerprotein alpha Globin, taking advantage of the fact that the globin genecontains introns as opposed to the intron-less histone genes. For thispurpose constructs were generated in which the alpha globin codingsequence was followed by a histone stem-loop signal (histone stem-loopfollowed by the histone downstream element) and a polyadenylation signal(Whitelaw, E., et al. (1986). Nucleic Acids Research, 14(17), 7059-7070;Pandey, N. B., & Marzluff, W. F. (1987). Molecular and Cellular Biology,7(12), 4557-4559; Pandey, N. B., et al. (1990). Nucleic Acids Research,18(11), 3161-3170).

In another approach Lüscher et al. investigated the cell-cycle dependentregulation of a recombinant histone H4 gene. Constructs were generatedin which the H4 coding sequence was followed by a histone stem-loopsignal and a polyadenylation signal, the two processing signalsincidentally separated by a galactokinase coding sequence (Lüscher, B.et al, (1985). Proc. Natl. Acad. Sci. USA, 82(13), 4389-4393).

Additionally, Stauber et al. identified the minimal sequence required toconfer cell-cycle regulation on histone H4 mRNA levels. For theseinvestigations constructs were used, comprising a coding sequence forthe selection marker Xanthine:guanine phosphoribosyl transferase (GPT)preceding a histone stem-loop signal followed by a polyadenylationsignal (Stauber, C. et al., (1986). EMBO J, 5(12), 3297-3303).

Examining histone pre-mRNA processing Wagner et al. identified factorsrequired for cleavage of histone pre-mRNAs using a reporter constructplacing EGFP between a histone stem-loop signal and a polyadenylationsignal, such that EGFP was expressed only in case histone pre-mRNAprocessing was disrupted (Wagner, E. J. et al., (2007). Mol Cell 28(4),692-9).

To be noted, translation of polyadenylated mRNA usually requires the 3′poly(A) sequence to be brought into proximity of the 5′ CAP. This ismediated through protein-protein interaction between the poly(A) bindingprotein and eukaryotic initiation factor eIF4G. With respect toreplication-dependent histone mRNAs, an analogous mechanism has beenuncovered. In this context, Gallie et al. show that the histonestem-loop is functionally similar to a poly(A) sequence in that itenhances translational efficiency and is co-dependent on a 5′-CAP inorder to establish an efficient level of translation. They showed thatthe histone stem-loop is sufficient and necessary to increase thetranslation of a reporter mRNA in transfected Chinese hamster ovarycells but must be positioned at the 3′-terminus in order to functionoptimally. Therefore, similar to the poly(A) tail on other mRNAs, the 3′end of these histone mRNAs appears to be essential for translation invivo and is functionally analogous to a poly(A) tail (Gallie, D. R.,Lewis, N. J., & Marzluff, W. F. (1996), Nucleic Acids Research, 24(10),1954-1962).

Additionally, it could be shown that SLBP is bound to the cytoplasmichistone mRNA and is required for its translation. Even though SLBP doesnot interact directly with eIF4G, the domain required for translation ofhistone mRNA interacts with the recently identified protein SLIP1. In afurther step, SLIP1 interacts with elF4G and allows to circularizehistone mRNA and to support efficient translation of histone mRNA by amechanism similar to the translation of polyadenylated mRNAs.

As mentioned above, gene therapy approaches normally use DNA to transferthe coding information into the cell which is then transcribed intomRNA, carrying the naturally occurring elements of an mRNA, particularlythe 5′-CAP structure and the 3′ poly(A) sequence to ensure expression ofthe encoded therapeutic or antigenic protein.

However, in many cases expression systems based on the introduction ofsuch nucleic acids into the patient's cells or tissue and the subsequentexpression of the desired polypeptides coded for by these nucleic acidsdo not exhibit the desired, or even the required, level of expressionwhich may allow for an efficient therapy, irrespective as to whether DNAor RNA is used.

In the prior art, different attempts have hitherto been made to increasethe yield of the expression of an encoded protein, in particular by useof improved expression systems, both in vitro and/or in vivo. Methodsfor increasing expression described generally in the prior art areconventionally based on the use of expression vectors or cassettescontaining specific promoters and corresponding regulation elements. Asthese expression vectors or cassettes are typically limited toparticular cell systems, these expression systems have to be adapted foruse in different cell systems. Such adapted expression vectors orcassettes are then usually transfected into the cells and typicallytreated in dependence of the specific cell line. Therefore, preferenceis given primarily to those nucleic acid molecules which are able toexpress the encoded proteins in a target cell by systems inherent in thecell, independent of promoters and regulation elements which arespecific for particular cell types. In this context, there can bedistinguished between mRNA stabilizing elements and elements whichincrease translation efficiency of the mRNA.

mRNAs which are optimized in their coding sequence and which are ingeneral suitable for such a purpose are described in application WO02/098443 (CureVac GmbH). For example, WO 02/098443 describes mRNAs thatare stabilised in general form and optimised for translation in theircoding regions. WO 02/098443 further discloses a method for determiningsequence modifications. WO 02/098443 additionally describespossibilities for substituting adenine and uracil nucleotides in mRNAsequences in order to increase the guanine/cytosine (G/C) content of thesequences. According to WO 02/098443, such substitutions and adaptationsfor increasing the G/C content can be used for gene therapeuticapplications but also genetic vaccines in the treatment of cancer orinfectious diseases. In this context, WO 02/098443 generally mentionssequences as a base sequence for such modifications, in which themodified mRNA codes for at least one biologically active peptide orpolypeptide, which is translated in the patient to be treated, forexample, either not at all or inadequately or with faults.Alternatively, WO 02/098443 proposes mRNAs coding for antigens e.g.allergenic antigens or auto-immune self-antigens or viral antigens as abase sequence for such modifications.

In a further approach to increase the expression of an encoded proteinthe application WO 2007/036366 describes the positive effect of longpoly(A) sequences (particularly longer than 120 bp) and the combinationof at least two 3′ untranslated regions of the beta globin gene on mRNAstability and translational activity.

However, even though all these latter prior art documents already try toprovide quite efficient tools for gene therapy approaches andadditionally improved mRNA stability and translational activity, therestill remains the problem of a generally lower stability of RNA-basedapplications versus DNA vaccines and DNA based gene therapeuticapproaches. Accordingly, there still exists a need in the art to provideimproved tools for gene therapy approaches and genetic vaccination or asa supplementary therapy for conventional treatments as discussed above,which allow for better provision of encoded proteins in vivo, e.g. via afurther improved mRNA stability and/or translational activity,preferably for gene therapy and genetic vaccination.

Furthermore despite of all progress in the art, efficient expression ofan encoded peptide or protein in cell-free systems, cells or organisms(recombinant expression) is still a challenging problem.

The object underlying the present invention is, therefore, to provideadditional and/or alternative methods to increase expression of anencoded protein, preferably via further stabilization of the mRNA and/oran increase of the translational efficiency of such an mRNA with respectto such nucleic acids known from the prior art for the use in geneticvaccination in the therapeutic or prophylactic treatment of allergies orautoimmune diseases.

This object is solved by the subject matter of the attached claims.Particularly, the object underlying the present invention is solvedaccording to a first aspect by an inventive nucleic acid sequencecomprising or coding for

-   -   a) a coding region, encoding at least one peptide or protein        which comprises an allergenic antigen or an autoimmune        self-antigen or a fragment, variant or derivative thereof;    -   b) at least one histone stem-loop, and    -   c) a poly(A) sequence or a polyadenylation signal, preferably        for increasing the expression of said encoded peptide or        protein.

Alternatively, any appropriate stem loop sequence other than a histonestem loop sequence (derived from histone genes, in particular fromhistone genes of the families H1, H2A, H2B, H3 and H4) may be employedby the present invention in all of its aspects and embodiments.

In this context it is particularly preferred that the inventive nucleicacid according to the first aspect of the present invention is producedat least partially by DNA or RNA synthesis, preferably as describedherein or is an isolated nucleic acid.

The present invention is based on the surprising finding of the presentinventors, that the combination of a poly(A) sequence or polyadenylationsignal and at least one histone stem-loop, even though both representingalternative mechanisms in nature, acts synergistically as thiscombination increases the protein expression manifold above the levelobserved with either of the individual elements. The synergistic effectof the combination of poly(A) and at least one histone stem-loop is seenirrespective of the order of poly(A) and histone stem-loop andirrespective of the length of the poly(A) sequence.

Therefore it is particularly preferred that the inventive nucleic acidsequence comprises or codes for a) a coding region, encoding at leastone peptide or protein which comprises an allergenic antigen or anautoimmune self-antigen or a fragment, variant or derivative thereof; b)at least one histone stem-loop, and c) a poly(A) sequence orpolyadenylation sequence; preferably for increasing the expression levelof said encoded peptide or protein. In some preferred embodiments, itmay be preferred if the encoded protein is not a histone protein, inparticular no histone protein of the H4, H3, H2A and/or H2B histonefamily or a fragment, derivative or variant thereof retaininghistone(-like) function), namely forming a nucleosome. Also, the encodedprotein typically does not correspond to a histone linker protein of theH1 histone family. The inventive nucleic acid molecule does typicallynot contain any regulatory signals (5′ and/or, particularly, 3′) of amouse histone gene, in particular not of a mouse histone gene H2A and,further, most preferably not of the mouse histone gene H2A614. Inparticular, it does not contain a histone stem loop and/or a histonestem loop processing signal from a mouse histone gene, in particular notof mouse histone gene H2A und, most preferably not of mouse histone geneH2A614.

Also, the inventive nucleic acid typically does not provide a reporterprotein (e.g. Luciferase, GFP, EGFP, β-Galactosidase, particularlyEGFP), galactokinase (galK) and/or marker or selection protein (e.g.alpha-Globin, Galactokinase and Xanthine:Guanine phosphoribosyltransferase (GPT)) or a bacterial reporter protein, e.g. chloramphenicolacetyl transferase (CAT) or other bacterial antibiotics resistanceproteins, e.g. derived from the bacterial neo gene in its element (a).

A reporter, marker or selection protein is typically understood not tobe a protein acting as an allergic antigen or autoimmune self-antigenaccording to the invention. A reporter, marker or selection protein orits underlying gene is commonly used as a research tool in bacteria,cell culture, animals or plants. They confer on organisms (preferablyheterologously) expressing them an easily identifiable property, whichmay be measured or which allows for selection. Specifically, marker orselection proteins exhibit a selectable function. Typically, suchselection, marker or reporter proteins do not naturally occur in humansor other mammals, but are derived from other organisms, in particularfrom bacteria or plants. Accordingly, proteins with selection, marker orreporter function originating from species other than mammals, inparticular other than humans, are preferably excluded from beingunderstood as “an allergic antigen or autoimmune self-antigen” accordingto the present invention. In particular, a selection, marker or reporterprotein allows to identify transformed cells by in vitro assays basede.g. on fluorescence or other spectroscopic techniques and resistancetowards antibiotics. Selection, reporter or marker genes awarding suchproperties to transformed cells are therefore typically not understoodto be an allergic antigen or autoimmune self-antigen according to theinvention.

In any case, reporter, marker or selection proteins do usually not exertany therapeutic effect by acting as an allergic antigen or autoimmuneself-antigen. If any single reporter, marker or selection protein shouldnevertheless do so (in addition to its reporter, selection or markerfunction), such a reporter, marker or selection protein is preferablynot understood to be a “an allergic antigen or autoimmune self-antigen”within the meaning of the present invention.

In contrast, an allergic antigen or autoimmune self-antigen (includingits fragments, variants and derivatives), in particular excludinghistone genes of the families H1, H2A, H2B, H3 and H4, according to thepresent invention does typically not exhibit a selection, marker orreporter function. If any single “an allergic antigen or autoimmuneself-antigen” nevertheless should do so (in addition to its therapeutic,allergic function), such a protein is preferably not understood to be a“selection, marker or reporter protein” within the meaning of thepresent invention.

It is most preferably understood that a an autoimmune self-antigenaccording to the invention is derived from mammals, in particularhumans, and does not qualify as selection, marker or reporter protein,while an allergic antigen is typically derived from plants or lowerorganisms, dogs, cats and does also not qualify as selection, marker orreporter protein.

Accordingly, it is preferred that the coding region (a) encoding atleast one peptide or protein is heterologous to at least (b) the atleast one histone stem loop, or more broadly, to any appropriate stemloop. In other words, “heterologous” in the context of the presentinvention means that the at least one stem loop sequence does notnaturally occur as a (regulatory) sequence (e.g. at the 3′UTR) of thespecific gene, which encodes the (allergic or autoimmune self-antigen)protein or peptide of element (a) of the inventive nucleic acid.Accordingly, the (histone) stem loop of the inventive nucleic acid isderived preferably from the 3′ UTR of a gene other than the onecomprising the coding region of element (a) of the inventive nucleicacid. E.g., the coding region of element (a) will not encode a histoneprotein or a fragment, variant or derivative thereof (retaining thefunction of a histone protein), if the inventive nucleic isheterologous, but will encode any other peptide or sequence (of the sameor another species) which exerts a biological function, preferablytherapeutic function (as a vaccine against these allergens andautoimmune self-antigens) other than a histone(-like) function, e.g.will encode an therapeutic protein (by exerting a therapeutic function,e.g. in terms vaccination of e.g. mammalian, in particular human,against allergic reactions triggered by exogenous allergens, e.g. plantallergens (e.g. birch etc.) or by autoimmune self-antigens.

In this context it is particularly preferred that the inventive nucleicacid comprises or codes for in 5′- to 3′-direction:

-   -   a) a coding region, encoding at least one peptide or protein        which comprises an allergenic antigen or an autoimmune        self-antigen or a fragment, variant or derivative thereof;    -   b) at least one histone stem-loop, optionally without a histone        downstream element (HDE) 3′ to the histone stem-loop    -   c) a poly(A) sequence or a polyadenylation signal.

The term “histone downstream element (HDE) refers to a purine-richpolynucleotide stretch of about 15 to 20 nucleotides 3′ of naturallyoccurring histone stem-loops, which represents the binding site for theU7 snRNA involved in processing of histone pre-mRNA into mature histonemRNA. For example in sea urchins the HDE is CAAGAAAGA (Dominski, Z. andW. F. Marzluff (2007), Gene 396(2): 373-90).

Furthermore it is preferable that the inventive nucleic acid accordingto the first aspect of the present invention does not comprise anintron.

In another particular preferred embodiment, the inventive nucleic acidsequence according to the first aspect of the present inventioncomprises or codes for from 5′ to 3′:

-   -   a) a coding region, preferably encoding at least one peptide or        protein which comprises an allergenic antigen or an autoimmune        self-antigen or a fragment, variant or derivative thereof;    -   c) a poly(A) sequence; and    -   b) at least one histone stem-loop.

The inventive nucleic acid sequence according to the first embodiment ofthe present invention comprise any suitable nucleic acid, selected e.g.from any (single-stranded or double-stranded) DNA, preferably, withoutbeing limited thereto, e.g. genomic DNA, plasmid DNA, single-strandedDNA molecules, double-stranded DNA molecules, or may be selected e.g.from any PNA (peptide nucleic acid) or may be selected e.g. from any(single-stranded or double-stranded) RNA, preferably a messenger RNA(mRNA); etc. The inventive nucleic acid sequence may also comprise aviral RNA (vRNA). However, the inventive nucleic acid sequence may notbe a viral RNA or may not contain a viral RNA. More specifically, theinventive nucleic acid sequence may not contain viral sequence elements,e.g. viral enhancers or viral promoters (e.g. no inactivated viralpromoter or sequence elements, more specifically not inactivated byreplacement strategies), or other viral sequence elements, or viral orretroviral nucleic acid sequences. More specifically, the inventivenucleic acid sequence may not be a retroviral or viral vector or amodified retroviral or viral vector.

In any case, the inventive nucleic acid sequence may or may not containan enhancer and/or promoter sequence, which may be modified or not orwhich may be activated or not. The enhancer and or promoter may be plantexpressible or not expressible, and/or in eukaryotes expressible or notexpressible and/or in prokaryotes expressible or not expressible. Theinventive nucleic acid sequence may contain a sequence encoding a(self-splicing) ribozyme or not.

In specific embodiments the inventive nucleic acid sequence may be ormay comprise a self-replicating RNA (replicon).

Preferably, the inventive nucleic acid sequence is a plasmid DNA, or anRNA, particularly an mRNA.

In particular embodiments of the first aspect of the present invention,the inventive nucleic acid is a nucleic acid sequence comprised in anucleic acid suitable for in vitro transcription, particularly in anappropriate in vitro transcription vector (e.g. a plasmid or a linearnucleic acid sequence comprising specific promoters for in vitrotranscription such as T3, T7 or Sp6 promoters).

In further particular preferred embodiments of the first aspect of thepresent invention, the inventive nucleic acid is comprised in a nucleicacid suitable for transcription and/or translation in an expressionsystem (e.g. in an expression vector or plasmid), particularly aprokaryotic (e.g. bacteria like E. coli) or eukaryotic (e.g. mammaliancells like CHO cells, yeast cells or insect cells or whole organismslike plants or animals) expression system.

The term “expression system” means a system (cell culture or wholeorganisms) which is suitable for production of peptides, proteins or RNAparticularly mRNA (recombinant expression).

The inventive nucleic acid sequence according to the first aspect of thepresent invention comprises or codes for at least one histone stem-loop.In the context of the present invention, such a histone stem-loop istypically derived from histone genes and comprises an intramolecularbase pairing of two neighbored entirely or partially reversecomplementary sequences, thereby forming a stem-loop. A stem-loop canoccur in single-stranded DNA or, more commonly, in RNA. The structure isalso known as a hairpin or hairpin loop and usually consists of a stemand a (terminal) loop within a consecutive sequence, wherein the stem isformed by two neighbored entirely or partially reverse complementarysequences separated by a short sequence as sort of spacer, which buildsthe loop of the stem-loop structure. The two neighbored entirely orpartially reverse complementary sequences may be defined as e.g. stemloop elements stem1 and stem2. The stem loop is formed when these twoneighbored entirely or partially reverse complementary sequences, e.g.stem loop elements stem1 and stem2, form base-pairs with each other,leading to a double stranded nucleic acid sequence comprising anunpaired loop at its terminal ending formed by the short sequencelocated between stem loop elements stem1 and stem2 on the consecutivesequence. The unpaired loop thereby typically represents a region of thenucleic acid which is not capable of base pairing with either of thesestem loop elements. The resulting lollipop-shaped structure is a keybuilding block of many RNA secondary structures. The formation of astem-loop structure is thus dependent on the stability of the resultingstem and loop regions, wherein the first prerequisite is typically thepresence of a sequence that can fold back on itself to form a paireddouble strand. The stability of paired stem loop elements is determinedby the length, the number of mismatches or bulges it contains (a smallnumber of mismatches is typically tolerable, especially for a longerdouble stranded stretch), and the base composition of the paired region.In the context of the present invention, a loop length of 3 to 15 basesis conceivable, while a more preferred optimal loop length is 3-10bases, more preferably 3 to 8, 3 to 7, 3 to 6 or even more preferably 4to 5 bases, and most preferably 4 bases. The stem sequence forming thedouble stranded structure typically has a length of between 5 to 10bases, more preferably, between 5 to 8 bases

In the context of the present invention, a histone stem-loop istypically derived from histone genes (e.g. genes from the histonefamilies H1, H2A, H2B, H3, H4) and comprises an intramolecular basepairing of two neighbored entirely or partially reverse complementarysequences, thereby forming a stem-loop. Typically, a histone 3′ UTRstem-loop is an RNA element involved in nucleocytoplasmic transport ofthe histone mRNAs, and in the regulation of stability and of translationefficiency in the cytoplasm. The mRNAs of metazoan histone genes lackpolyadenylation and a poly-A tail, instead 3′ end processing occurs at asite between this highly conserved stem-loop and a purine rich regionaround 20 nucleotides downstream (the histone downstream element, orHDE). The histone stem-loop is bound by a 31 kDa stem-loop bindingprotein (SLBP—also termed the histone hairpin binding protein, or HBP).Such histone stem-loop structures are preferably employed by the presentinvention in combination with other sequence elements and structures,which do not occur naturally (which means in untransformed livingorganisms/cells) in histone genes, but are combined—according to theinvention—to provide an artificial, heterologous nucleic acid.Accordingly, the present invention is particularly based on the findingthat an artificial (non-native) combination of a histone stem-loopstructure with other heterologous sequence elements, which do not occurin histone genes or metazoan histone genes and are isolated fromoperational and/or regulatory sequence regions (influencingtranscription and/or translation) of genes coding for proteins otherthan histones, provide advantageous effects. Accordingly, one aspect ofthe invention provides the combination of a histone stem-loop structurewith a poly(A) sequence or a sequence representing a polyadenylationsignal (3′-terminal of a coding region), which does not occur inmetazoan histone genes.

According to another preferred aspect of the invention, a combination ofa histone stem-loop structure with a coding region coding for a protein(acting as an allergen or acting as an autoimmune self-antigen), whichdoes, preferably not occur in metazoan histone genes, is providedherewith (coding region and histone stem loop sequence areheterologous). It is preferred, if an autuimmine self-antigen occurs asa protein naturally in mammalians, preferably humans. In a still furtherpreferred embodiment, all the elements (a), (b) and (c) of the inventivenucleic acid are heterologous to each other and are combinedartificially from three different sources, e.g. (a) the autoimmuneself-antigen coding region from a human gene or the allergen fromanother source, in pasrticular a non-human organism (e.g. a plant, a petetc.) (b) the histone stem loop from an untranlated region of ametazoan, e.g. mammalian, non-human or human, histone gene and (c) thepoly(A) sequence or the polyadenylation signal from e.g. an untranlatedregion of a gene other than a histone gene and other than the genecoding for the allergen or the autoimmune self-antigen according toelement (a) of such an inventive nucleic acid.

A histone stem loop is, therefore, a stem-loop structure as describedherein, which, if preferably functionally defined, exhibits/retains theproperty of binding to its natural binding partner, the stem-loopbinding protein (SLBP—also termed the histone hairpin binding protein,or HBP).

According to the present invention the histone stem loop sequenceaccording to component (b) of claim 1 may not derived from a mousehistone protein. More specifically, the histone stem loop sequence maynot be derived from mouse histone gene H2A614. Also, the nucleic acid ofthe invention may neither contain a mouse histone stem loop sequence norcontain mouse histone gene H2A614. Further, the inventive nucleic acidsequence may not contain a stem-loop processing signal, morespecifically, a mouse histone processing signal and, most specifically,may not contain mouse stem loop processing signal H2kA614, even if theinventive nucleic acid sequence may contain at least one mammalianhistone gene. However, the at least one mammalian histone gene may notbe Seq. ID No. 7 of WO 01/12824.

According to one preferred embodiment of the first inventive aspect, theinventive nucleic acid sequence comprises or codes for at least onehistone stem-loop sequence, preferably according to at least one of thefollowing formulae (I) or (II):

Formula (I) (Stem-Loop Sequence without Stem Bordering Elements):

Formula (II) (Stem-Loop Sequence with Stem Bordering Elements):

wherein:

-   stem1 or stem2 bordering elements N₁₋₆ is a consecutive sequence of    1 to 6, preferably of 2 to 6, more preferably of 2 to 5, even more    preferably of 3 to 5, most preferably of 4 to 5 or 5 N, wherein each    N is independently from another selected from a nucleotide selected    from A, U, T, G and C, or a nucleotide analogue thereof;-   stem1 [N₀₋₂GN₃₋₅] is reverse complementary or partially reverse    complementary with element stem2, and is a consecutive sequence    between of 5 to 7 nucleotides;    -   wherein N₀₋₂ is a consecutive sequence of 0 to 2, preferably of        0 to 1, more preferably of 1 N, wherein each N is independently        from another selected from a nucleotide selected from A, U, T, G        and C or a nucleotide analogue thereof;    -   wherein N₃₋₅ is a consecutive sequence of 3 to 5, preferably of        4 to 5, more preferably of 4 N, wherein each N is independently        from another selected from a nucleotide selected from A, U, T, G        and C or a nucleotide analogue thereof, and    -   wherein G is guanosine or an analogue thereof, and may be        optionally replaced by a cytidine or an analogue thereof,        provided that its complementary nucleotide cytidine in stem2 is        replaced by guanosine;-   loop sequence [N₀₋₄(U/T)N₀₋₄] is located between elements stem1 and    stem2, and is a consecutive sequence of 3 to 5 nucleotides, more    preferably of 4 nucleotides;    -   wherein each N₀₋₄ is independent from another a consecutive        sequence of 0 to 4, preferably of 1 to 3, more preferably of 1        to 2 N, wherein each N is independently from another selected        from a nucleotide selected from A, U, T, G and C or a nucleotide        analogue thereof; and    -   wherein U/T represents uridine, or optionally thymidine;-   stem2 [N₃₋₅CN₀₋₂] is reverse complementary or partially reverse    complementary with element stem1, and is a consecutive sequence    between of 5 to 7 nucleotides;    -   wherein N₃₋₅ is a consecutive sequence of 3 to 5, preferably of        4 to 5, more preferably of 4 N, wherein each N is independently        from another selected from a nucleotide selected from A, U, T, G        and C or a nucleotide analogue thereof;    -   wherein N₀₋₂ is a consecutive sequence of 0 to 2, preferably of        0 to 1, more preferably of 1 N, wherein each N is independently        from another selected from a nucleotide selected from A, U, T, G        or C or a nucleotide analogue thereof; and    -   wherein C is cytidine or an analogue thereof, and may be        optionally replaced by a guanosine or an analogue thereof        provided that its complementary nucleotide guanosine in stem1 is        replaced by cytidine;    -   wherein    -   stem1 and stem2 are capable of base pairing with each other        forming a reverse complementary sequence, wherein base pairing        may occur between stem1 and stem2, e.g. by Watson-Crick base        pairing of nucleotides A and U/T or G and C or by        non-Watson-Crick base pairing e.g. wobble base pairing, reverse        Watson-Crick base pairing, Hoogsteen base pairing, reverse        Hoogsteen base pairing or are capable of base pairing with each        other forming a partially reverse complementary sequence,        wherein an incomplete base pairing may occur between stem1 and        stem2, on the basis that one or more bases in one stem do not        have a complementary base in the reverse complementary sequence        of the other stem.

In the above context, a wobble base pairing is typically anon-Watson-Crick base pairing between two nucleotides. The four mainwobble base pairs in the present context, which may be used, areguanosine-uridine, inosine-uridine, inosine-adenosine, inosine-cytidine(G-U/T, I-U/T, I-A and I-C) and adenosine-cytidine (A-C).

Accordingly, in the context of the present invention, a wobble base is abase, which forms a wobble base pair with a further base as describedabove. Therefore non-Watson-Crick base pairing, e.g. wobble basepairing, may occur in the stem of the histone stem-loop structureaccording to the present invention.

In the above context a partially reverse complementary sequencecomprises maximally 2, preferably only one mismatch in thestem-structure of the stem-loop sequence formed by base pairing of stem1and stem2. In other words, stem1 and stem2 are preferably capable of(full) base pairing with each other throughout the entire sequence ofstem1 and stem2 (100% of possible correct Watson-Crick ornon-Watson-Crick base pairings), thereby forming a reverse complementarysequence, wherein each base has its correct Watson-Crick ornon-Watson-Crick base pendant as a complementary binding partner.Alternatively, stem1 and stem2 are preferably capable of partial basepairing with each other throughout the entire sequence of stem1 andstem2, wherein at least about 70%, 75%, 80%, 85%, 90%, or 95% of the100% possible correct Watson-Crick or non-Watson-Crick base pairings areoccupied with the correct Watson-Crick or non-Watson-Crick base pairingsand at most about 30%, 25%, 20%, 15%, 10%, or 5% of the remaining basesare unpaired.

According to a preferred embodiment of the first inventive aspect, theat least one histone stem-loop sequence (with stem bordering elements)of the inventive nucleic acid sequence as defined herein comprises alength of about 15 to about 45 nucleotides, preferably a length of about15 to about 40 nucleotides, preferably a length of about 15 to about 35nucleotides, preferably a length of about 15 to about 30 nucleotides andeven more preferably a length of about 20 to about 30 and mostpreferably a length of about 24 to about 28 nucleotides.

According to a further preferred embodiment of the first inventiveaspect, the at least one histone stem-loop sequence (without stembordering elements) of the inventive nucleic acid sequence as definedherein comprises a length of about 10 to about 30 nucleotides,preferably a length of about 10 to about 20 nucleotides, preferably alength of about 12 to about 20 nucleotides, preferably a length of about14 to about 20 nucleotides and even more preferably a length of about 16to about 17 and most preferably a length of about 16 nucleotides.

According to a further preferred embodiment of the first inventiveaspect, the inventive nucleic acid sequence according to the firstaspect of the present invention may comprise or code for at least onehistone stem-loop sequence according to at least one of the followingspecific formulae (Ia) or (IIa):

Formula (Ia) (Stem-Loop Sequence without Stem Bordering Elements):

Formula (IIa) (Stem-Loop Sequence with Stem Bordering Elements):

wherein:N, C, G, T and U are as defined above.

According to a further more particularly preferred embodiment of thefirst aspect, the inventive nucleic acid sequence may comprise or codefor at least one histone stem-loop sequence according to at least one ofthe following specific formulae (Ib) or (IIb):

Formula (Ib) (Stem-Loop Sequence without Stem Bordering Elements):

Formula (IIb) (Stem-Loop Sequence with Stem Bordering Elements):

wherein:N, C, G, T and U are as defined above.

According to an even more preferred embodiment of the first inventiveaspect, the inventive nucleic acid sequence according to the firstaspect of the present invention may comprise or code for at least onehistone stem-loop sequence according to at least one of the followingspecific formulae (Ic) to (Ih) or (IIc) to (IIh), shown alternatively inits stem-loop structure and as a linear sequence representing histonestem-loop sequences as generated according to Example 1:

Formula (Ic): (Metazoan and Protozoan Histone Stem-Loop ConsensusSequence without Stem Bordering Elements):

(stem-loop structure) N U N   N N-N N-N N-N N-N G-C N-N(linear sequence) (SEQ ID NO: 1) NGNNNNNNUNNNNNCNFormula (IIc): (Metazoan and Protozoan Histone Stem-Loop ConsensusSequence with Stem Bordering Elements):

(stem-loop structure) N U N   N N-N N-N N-N N-N G-C  N*N*NNNN-NNNN*N*N*(linear sequence) (SEQ ID NO: 2) N*N*NNNNGNNNNNNUNNNNNCNNNN*N*N*Formula (Id): (without Stem Bordering Elements)

(stem-loop structure) N U N   N N-N N-N N-N N-N C-G N-N(linear sequence) (SEQ ID NO: 3) NCNNNNNNUNNNNNGNFormula (IId): (with Stem Bordering Elements)

(stem-loop structure) N U N   N N-N N-N N-N N-N C-G  N*N*NNNN-NNNN*N*N*(linear sequence) (SEQ ID NO: 4) N*N*NNNNCNNNNNNUNNNNNGNNNN*N*N*Formula (Ie): (Protozoan Histone Stem-Loop Consensus Sequence withoutStem Bordering Elements)

(stem-loop structure) N U N   N N-N N-N N-N N-N G-C D-H(linear sequence) (SEQ ID NO: 5) DGNNNNNNUNNNNNCHFormula (IIe): (Protozoan Histone Stem-Loop Consensus Sequence with StemBordering Elements)

(stem-loop structure) N U N   N N-N N-N N-N N-N G-C  N*N*NNND-HNNN*N*N*(linear sequence) (SEQ ID NO: 6) N*N*NNNDGNNNNNNUNNNNNCHNNN*N*N*Formula (If): (Metazoan Histone Stem-Loop Consensus Sequence withoutStem Bordering Elements)

(stem-loop structure) N U N   N Y-V Y-N B-D N-N G-C N-N(linear sequence) (SEQ ID NO: 7) NGNBYYNNUNVNDNCNFormula (IIf): (Metazoan Histone Stem-Loop Consensus Sequence with StemBordering Elements)

(stem-loop structure) N U N   N Y-V Y-N B-D N-N G-C  N*N*NNNN-NNNN*N*N*(linear sequence) (SEQ ID NO: 8) N*N*NNNNGNBYYNNUNVNDNCNNNN*N*N*Formula (Ig): (Vertebrate Histone Stem-Loop Consensus Sequence withoutStem Bordering Elements)

(stem-loop structure) N U D   H Y-A Y-B Y-R H-D G-C N-N(linear sequence) (SEQ ID NO: 9) NGHYYYDNUHABRDCNFormula (IIg): (Vertebrate Histone Stem-Loop Consensus Sequence withStem Bordering Elements)

(stem-loop structure) N U D   H Y-A Y-B Y-R H-D G-C  N*N*HNNN-NNNN*N*H*(linear sequence) (SEQ ID NO: 10) N*N*HNNNGHYYYDNUHABRDCNNNN*N*H*Formula (Ih): (Human Histone Stem-Loop Consensus Sequence (Homo sapiens)without Stem Bordering Elements)

(stem-loop structure) Y U D   H U-A C-S Y-R H-R G-C D-C(linear sequence) (SEQ ID NO: 11) DGHYCUDYUHASRRCCFormula (IIh): (Human Histone Stem-Loop Consensus Sequence (Homosapiens) with Stem Bordering Elements)

(stem loop structure) Y U D   H U-A C-S Y-R H-R G-C  N*H*AAHD-CVHB*N*H*(linear sequence) (SEQ ID NO: 12) N*H*AAHDGHYCUDYUHASRRCCVHB*N*H*wherein in each of above formulae (Ic) to (Ih) or (IIc) to (IIh):N, C, G, A, T and U are as defined above;each U may be replaced by T;each (highly) conserved G or C in the stem elements 1 and 2 may bereplaced by its complementary nucleotide base C or G, provided that itscomplementary nucleotide in the corresponding stem is replaced by itscomplementary nucleotide in parallel; and/orG, A, T, U, C, R, Y, M, K, S, W, H, B, V, D, and N are nucleotide basesas defined in the following Table:

abbreviation Nucleotide bases remark G G Guanine A A Adenine T T ThymineU U Uracile C C Cytosine R G or A Purine Y T/U or C Pyrimidine M A or CAmino K G or T/U Keto S G or C Strong (3H bonds) W A or T/U Weak (2Hbonds) H A or C or T/U Not G B G or T/U or C Not A V G or C or A Not T/UD G or A or T/U Not C N G or C or T/U or A Any base * Present or notBase may be present or not

In this context it is particularly preferred that the histone stem-loopsequence according to at least one of the formulae (I) or (la) to (Ih)or (II) or (IIa) to (IIh) of the present invention is selected from anaturally occurring histone stem loop sequence, more particularlypreferred from protozoan or metazoan histone stem-loop sequences, andeven more particularly preferred from vertebrate and mostly preferredfrom mammalian histone stem-loop sequences especially from human histonestem-loop sequences.

According to a particularly preferred embodiment of the first aspect,the histone stem-loop sequence according to at least one of the specificformulae (I) or (la) to (Ih) or (II) or (IIa) to (IIh) of the presentinvention is a histone stem-loop sequence comprising at each nucleotideposition the most frequently occurring nucleotide, or either the mostfrequently or the second-most frequently occurring nucleotide ofnaturally occurring histone stem-loop sequences in metazoa and protozoa(FIG. 1), protozoa (FIG. 2), metazoa (FIG. 3), vertebrates (FIG. 4) andhumans (FIG. 5) as shown in FIG. 1-5. In this context it is particularlypreferred that at least 80%, preferably at least 85%, or most preferablyat least 90% of all nucleotides correspond to the most frequentlyoccurring nucleotide of naturally occurring histone stem-loop sequences.

In a further particular embodiment of the first aspect, the histonestem-loop sequence according to at least one of the specific formulae(I) or (la) to (Ih) of the present invention is selected from followinghistone stem-loop sequences (without stem-bordering elements)representing histone stem-loop sequences as generated according toExample 1:

(SEQ ID NO: 13 according to formula (Ic)) VGYYYYHHTHRVVRCB(SEQ ID NO: 14 according to formula (Ic)) SGYYYTTYTMARRRCS(SEQ ID NO: 15 according to formula (Ic)) SGYYCTTTTMAGRRCS(SEQ ID NO: 16 according to formula (Ie)) DGNNNBNNTHVNNNCH(SEQ ID NO: 17 according to formula (Ie)) RGNNNYHBTHRDNNCY(SEQ ID NO: 18 according to formula (Ie)) RGNDBYHYTHRDHNCY(SEQ ID NO: 19 according to formula (If)) VGYYYTYHTHRVRRCB(SEQ ID NO: 20 according to formula (If)) SGYYCTTYTMAGRRCS(SEQ ID NO: 21 according to formula (If)) SGYYCTTTTMAGRRCS(SEQ ID NO: 22 according to formula (Ig)) GGYYCTTYTHAGRRCC(SEQ ID NO: 23 according to formula (Ig)) GGCYCTTYTMAGRGCC(SEQ ID NO: 24 according to formula (Ig)) GGCTCTTTTMAGRGCC(SEQ ID NO: 25 according to formula (Ih)) DGHYCTDYTHASRRCC(SEQ ID NO: 26 according to formula (Ih)) GGCYCTTTTHAGRGCC(SEQ ID NO: 27 according to formula (Ih)) GGCYCTTTTMAGRGCC

Furthermore in this context following histone stem-loop sequences (withstem bordering elements) as generated according to Example 1 accordingto one of specific formulae (II) or (IIa) to (IIh) are particularlypreferred:

(SEQ ID NO: 28 according to formula (IIc))H*H*HHVVGYYYYHHTHRVVRCBVHH*N*N*(SEQ ID NO: 29 according to formula (IIc))M*H*MHMSGYYYTTYTMARRRCSMCH*H*H*(SEQ ID NO: 30 according to formula (IIc))M*M*MMMSGYYCTTTTMAGRRCSACH*M*H*(SEQ ID NO: 31 according to formula (IIe))N*N*NNNDGNNNBNNTHVNNNCHNHN*N*N*(SEQ ID NO: 32 according to formula (IIe))N*N*HHNRGNNNYHBTHRDNNCYDHH*N*N*(SEQ ID NO: 33 according to formula (IIe))N*H*HHVRGNDBYHYTHRDHNCYRHH*H*H*(SEQ ID NO: 34 according to formula (IIf))H*H*MHMVGYYYTYHTHRVRRCBVMH*H*N*(SEQ ID NO: 35 according to formula (IIf))M*M*MMMSGYYCTTYTMAGRRCSMCH*H*H*(SEQ ID NO: 36 according to formula (IIf))M*M*MMMSGYYCTTTTMAGRRCSACH*M*H*(SEQ ID NO: 37 according to formula (IIg))H*H*MAMGGYYCTTYTHAGRRCCVHN*N*M*(SEQ ID NO: 38 according to formula (IIg))H*H*AAMGGCYCTTYTMAGRGCCVCH*H*M*(SEQ ID NO: 39 according to formula (IIg))M*M*AAMGGCTCTTTTMAGRGCCMCY*M*M*(SEQ ID NO: 40 according to formula (IIh))N*H*AAHDGHYCTDYTHASRRCCVHB*N*H*(SEQ ID NO: 41 according to formula (IIh))H*H*AAMGGCYCTTTTHAGRGCCVMY*N*M*(SEQ ID NO: 42 according to formula (IIh))H*M*AAAGGCYCTTTTMAGRGCCRMY*H*M*

According to a further preferred embodiment of the first inventiveaspect, the inventive nucleic acid sequence comprises or codes for atleast one histone stem-loop sequence showing at least about 80%,preferably at least about 85%, more preferably at least about 90%, oreven more preferably at least about 95%, sequence identity with the notto 100% conserved nucleotides in the histone stem-loop sequencesaccording to at least one of specific formulae (I) or (Ia) to (Ih) or(II) or (IIa) to (IIh) or with a naturally occurring histone stem-loopsequence.

In a preferred embodiment, the histone stem loop sequence does notcontain the loop sequence 5′-UUUC-3′. More specifically, the histonestem loop does not contain the stem1 sequence 5′-GGCUCU-3′ and/or thestem2 sequence 5′-AGAGCC-3′, respectively. In another preferredembodiment, the stem loop sequence does not contain the loop sequence5′-CCUGCCC-3′ or the loop sequence 5′-UGAAU-3′. More specifically, thestem loop does not contain the stem1 sequence 5′-CCUGAGC-3′ or does notcontain the stem1 sequence 5′-ACCUUUCUCCA-3′ and/or the stem2 sequence5′-GCUCAGG-3′ or 5′-UGGAGAAAGGU-3′, respectively. Also, as far as theinvention is not limited to histone stem loop sequences specifically,stem loop sequences are preferably not derived from a mammalian insulinreceptor 3′-untranslated region. Also, preferably, the inventive nucleicacid may not contain histone stem loop processing signals, in particularnot those derived from mouse histone gene H2A614 gene (H2kA614).

The inventive nucleic acid sequence according to the first aspect of thepresent invention may optionally comprise or code for a poly(A)sequence. When present, such a poly(A) sequence comprises a sequence ofabout 25 to about 400 adenosine nucleotides, preferably a sequence ofabout 30 or, more preferably, of about 50 to about 400 adenosinenucleotides, more preferably a sequence of about 50 to about 300adenosine nucleotides, even more preferably a sequence of about 50 toabout 250 adenosine nucleotides, most preferably a sequence of about 60to about 250 adenosine nucleotides. In this context the term “about”refers to a deviation of ±10% of the value(s) it is attached to.Accordingly, the poly(A) sequence contains at least 25 or more than 25,more preferably, at least 30, more preferably at least 50 adenosinenucleotides. Therefore, such a poly (A) sequence does typically notcontain less than 20 adenosine nucleotides. More particularly, it doesnot contain 10 and/or less than 10 adenosine nucleotides.

Preferably, the nucleic acid according of the present invention does notcontain one or two or at least one or all but one or all of thecomponents of the group consisting of: a sequence encoding a ribozyme(preferably a self-splicing ribozyme), a viral nucleic acid sequence, ahistone stem-loop processing signal, in particular a histone-stem loopprocessing sequence derived from mouse histone H2A614 gene, a Neo gene,an inactivated promoter sequence and an inactivated enhancer sequence.Even more preferably, the nucleic acid according to the invention doesnot contain a ribozyme, preferably a self-splicing ribozyme, and one ofthe group consisting of: a Neo gene, an inactivated promoter sequence,an inactivated enhancer sequence, a histone stem-loop processing signal,in particular a histone-stem loop processing sequence derived from mousehistone H2A614 gene. Accordingly, the nucleic acid may in a preferredmode neither contain a ribozyme, preferably a self-splicing ribozyme,nor a Neo gene or, alternatively, neither a ribozyme, preferably aself-splicing ribozyme, nor any resistance gene (e.g. usually appliedfor selection). In another preferred mode, the nucleic acid of theinvention may neither contain a ribozyme, preferably a self-splicingribozyme nor a histone stem-loop processing signal, in particular ahistone-stem loop processing sequence derived from mouse histone H2A614gene.

Alternatively, according to the first aspect of the present invention,the inventive nucleic sequence optionally comprises a polyadenylationsignal which is defined herein as a signal which conveys polyadenylationto a (transcribed) mRNA by specific protein factors (e.g. cleavage andpolyadenylation specificity factor (CPSF), cleavage stimulation factor(CstF), cleavage factors I and II (CF I and CF II), poly(A) polymerase(PAP)). In this context a consensus polyadenylation signal is preferredcomprising the NN(U/T)ANA consensus sequence. In a particular preferredaspect the polyadenylation signal comprises one of the followingsequences: AA(U/T)AAA or A(U/T)(UIT)AAA (wherein uridine is usuallypresent in RNA and thymidine is usually present in DNA). In someembodiments, the polyadenylation signal used in the inventive nucleicacid does not correspond to the U3 snRNA, U5, the polyadenylationprocessing signal from human gene G-CSF, or the SV40 polyadenylationsignal sequences. In particular, the above polyadenylation signals arenot combined with any antibiotics resistance gene (or any otherreporter, marker or selection gene), in particular not with theresistance neo gene (neomycin phosphotransferase) (as the gene of thecoding region according to element (a) of the inventive nucleic acid.And, any of the above polyadenylation signals are preferably notcombined with the histone stem loop or the histone stem loop processingsignal from mouse histone gene H2A614 in an inventive nucleic acid.

The inventive nucleic acid sequence according to the first aspect of thepresent invention furthermore encodes a protein or a peptide, whichcomprises an allergenic antigen or an autoimmune self-antigen or afragment, variant or derivative thereof.

Such allergenic antigens may be selected from antigens derived fromdifferent sources, e.g. from animals, plants, fungi, bacteria, etc.Sources of allergens in this context include e.g. grasses, pollens,molds, drugs, or numerous environmental triggers, etc. Allergenicantigens typically belong to different classes of compounds, such asnucleic acids and their fragments, proteins or peptides and theirfragments, carbohydrates, polysaccharides, sugars, lipids,phospholipids, etc. Of particular interest in the context of the presentinvention are protein or peptide antigens and their fragments orepitopes, or nucleic acids and their fragments, particularly nucleicacids and their fragments, encoding such protein or peptide antigens andtheir fragments or epitopes.

Antigens associated with allergy or allergic disease (allergens orallergenic antigens) are preferably derived from a source selected fromthe list consisting of:

Acarus spp (Aca s 1, Aca s 10, Aca s 10.0101, Aca s 13, Aca s 13.0101,Aca s 2, Aca s 3, Aca s 7, Aca s 8), Acanthocybium spp (Aca so 1),Acanthocheilonema spp (Aca v 3, Aca v 3.0101), Acetes spp (Ace ja 1),Actinidia spp (Act a 1, Act c 1, Act c 10, Act c 10.0101, Act c 2, Act c4, Act c 5, Act c 5.0101, Act c 8, Act c 8.0101, Act c Chitinase, Act d1, Act d 1.0101, Act d 10, Act d 10.0101, Act d 10.0201, Act d 11, Act d11.0101, Act d 2, Act d 2.0101, Act d 3, Act d 3.0101, Act d 3.02, Act d4, Act d 4.0101, Act d 5, Act d 5.0101, Act d 6, Act d 6.0101, Act d 7,Act d 7.0101, Act d 8, Act d 8.0101, Act d 9, Act d 9.0101, Act dChitinase, Act e 1, Act e 5), Acyrthosiphon spp (Acy pi 7, Acy pi7.0101, Acy pi 7.0102), Adenia spp (Ade v RIP), Aedes spp (Aed a 1, Aeda 1.0101, Aed a 2, Aed a 2.0101, Aed a 3, Aed a 3.0101, Aed a 4, Aed a7, Aed a 7.0101, Aed a 7.0102, Aed a 7.0103, Aed a 7.0104, Aed a 7.0105,Aed a 7.0106, Aed a 7.0107, Aed a 7.0108, Aed a 7.0109, Aed a 7.0110,Aed a 7.0111, Aed al 1, Aed al 3, Aed al 37kD, Aed v 37kD, Aed v 63kD),Aegilops spp (Aeg ta 28, Aeg ta alpha_Gliadin, Aeg um 28, Aeg un 28),Aethaloperca spp (Aet ro 1), Agropyron spp (Agr c 7), Agrostis spp (Agrca 1, Agr ca 5, Agr g 1, Agr g 4, Agr s 5), Agrobacterium spp (Agr spCP4 EPSPS), Ailuropoda spp (Ail me Phosvitin, Ail me TCTP), Aix spp (Aixga 1, Aix sp 1), Aleuroglyphus spp (Ale o 1, Ale o 10, Ale o 10.0101,Ale o 10.0102, Ale o 13, Ale o 14, Ale o 2, Ale o 20, Ale o 3, Ale o 5,Ale o 7, Ale o 8, Ale o 9), Allium spp (All a 3, All a Alliin lyase, Allc 3, All c 30kD, All c 4, All c Alliin lyase, All p Alliin lyase, All sAlliin lyase), Alnus spp (Aln g 1, AIn g 1.0101, Aln g 1/Bet v 1/Cor a 1TPC7, AIn g 1/Bet v 1/Cor a 1 TPC9, AIn g 2, Aln g 4, AIn g 4.0101),Alopochen spp (Alo ae 1), Alopecurus spp (Alo p 1, Alo p 5), Alternariaspp (Alt a 1, Alt a 1.0101, Alt a 1.0102, Alt a 10, Alt a 10.0101, Alt a12, Alt a 12.0101, Alt a 13, Alt a 13.0101, Alt a 2, Alt a 3, Alt a3.0101, Alt a 4, Alt a 4.0101, Alt a 5, Alt a 5.0101, Alt a 6, Alt a6.0101, Alt a 7, Alt a 7.0101, Alt a 70kD, Alt a 8, Alt a 8.0101, Alt a9, Alt a MnSOD, Alt a NTF2, Alt a TCTP, Alt ar 1, Alt arg 1, Alt b 1,Alt bI 1, Alt br 1, Alt c 1, Alt ca 1, Alt ce 1, Alt ch 1, Alt ci 1, Altco 1, Alt cr 1, Alt ct 1, Alt cu 1, Alt cy 1, Alt d 1, Alt du 1, Alt e1, Alt et 1, Alt eu 1, Alt ga 1, Alt gr 1, Alt j 1, Alt 11, Alt lo 1,Alt m 1, Alt me 1, Alt mi 1, Alt mo 1, Alt o 1, Alt p 1, Alt ph 1, Altpo 1, Alt ps 1, Alt r 1, Alt s 1, Alt se 1, Alt sm 1, Alt so 1, Alt su1, Alt t 1, Alt to 1, Alt to 1), Amaranthus spp (Ama r 2, Ama r 2.0101,Ama v 2, Ama v 2.0101, Ama v 2.0201), Ambrosia spp (Amb a 1, Amb a1.0101, Amb a 1.0201, Amb a 1.0202, Amb a 1.0301, Amb a 1.0302, Amb a1.0303, Amb a 1.0304, Amb a 1.0305, Amb a 1.0401, Amb a 1.0402, Amb a1.0501, Amb a 1.0502, Amb a 10, Amb a 10.0101, Amb a 3, Amb a 3.0101,Amb a 4, Amb a 4.0101, Amb a 5, Amb a 5.0101, Amb a 6, Amb a 6.0101, Amba 7, Amb a 7.0101, Amb a 8, Amb a 8.0101, Amb a 8.0102, Amb a 9, Amb a9.0101, Amb a 9.0102, Amb a CPI, Amb p 1, Amb p 5, Amb p 5.0101, Amb p5.0201, Amb t 5, Amb t 5.0101, Amb t 8), Ammothea spp (Amm h 7, Amm h7.0101), Anadara spp (Ana br 1), Ananas spp (Ana c 1, Ana c 1.0101, Anac 2, Ana c 2.0101, Ana c 2.0101 (MUXF3)), Anas spp (Ana ca 1),Anarhichas spp (Ana I 1), Anacardium spp (Ana o 1, Ana o 1.0101, Ana o1.0102, Ana o 2, Ana o 2.0101, Ana o 3, Ana o 3.0101), Anas spp (Ana p1, Ana p 2, Ana p 3), Anguilla spp (Ang a 1, Ang j 1), Anisakis spp (Anis 1, Ani s 1.0101, Ani s 10, Ani s 10.0101, Ani s 11, Ani s 11.0101, Anis 12, Ani s 12.0101, Ani s2, Ani s 2.0101, Ani s 24kD, Ani s 3, Ani s3.0101, Ani s 4, Ani s 4.0101, Ani s 5, Ani s 5.0101, Ani s 6, Ani s6.0101, Ani s 7, Ani s 7.0101, Ani s 8, Ani s 8.0101, Ani s 9, Ani s9.0101, Ani s CCOS3, Ani s Cytochrome B, Ani s FBPP, Ani s NADHDS4L, Anis NARaS, Ani s PEPB, Ani s Troponin), Annona spp (Ann c Chitinase),Anopheles spp (Ano da 17, Ano da 17.0101, Ano da 27, Ano da 27.0101, Anoda 7, Ano da 7.0101, Ano g 7, Ano g 7.0101), Anser spp (Ans a 1, Ans a2, Ans a 3, Ans in 1), Anthoxanthum spp (Ant o 1, Ant o 1.0101, Ant o12, Ant o 13, Ant o 2, Ant o 4, Ant o 5, Ant o 6, Ant o 7), Apis spp(Api c 1, Api c 1.0101, Api c 10, Api c 2, Api c 4, Api d 1, Api d1.0101, Api d 4, Api fl 4), Apium spp (Api g 1, Api g 1.0101, Api g1.0201, Api g 2, Api g 2.0101, Api g 3, Api g 3.0101, Api g 4, Api g4.0101, Api g 5, Api g 5.0101, Api g 6, Api g 6.0101), Apis spp (Api m1, Api m 1.0101, Api m 10, Api m 10.0101, Api m 11, Api m 11.0101, Api m11.0201, Api m 13kD, Api m 2, Api m 2.0101, Api m 3, Api m 3.0101, Api m4, Api m 4.0101, Api m 5, Api m 5.0101, Api m 6, Api m 6.0101, Api m 7,Api m 7.0101, Api m 8, Api m 8.0101, Api m 9, Api m 9.0101, Api m A1-A2,Api m A1-A2-A3, Api m Apalbumin 1, Api m Apalbumin 2, Api me 1, Api me4), Arachis spp (Ara d 2, Ara d 6, Ara f 3, Ara f 4, Ara h 1, Ara h1.0101, Ara h 10, Ara h 10.0101, Ara h 10.0102, Ara h 11, Ara h 11.0101,Ara h 2, Ara h 2.0101, Ara h 2.0102, Ara h 2.0201, Ara h 2.0202, Ara h3, Ara h 3.0101, Ara h 4, Ara h 4.0101, Ara h 5, Ara h 5.0101, Ara h 6,Ara h 6.0101, Ara h 7, Ara h 7.0101, Ara h 7.0201, Ara h 7.0202, Ara h8, Ara h 8.0101, Ara h 8.0201, Ara h 9, Ara h 9.0101, Ara h 9.0201, Arah Agglutinin, Ara h Oleosin 18kD, Ara i 2, Ara i 6), Arabidopsis spp(Ara t 3, Ara t 8, Ara t GLP), Archosargus spp (Arc pr 1),Archaeopotamobius spp (Arc s 8, Arc s 8.0101), Aequipecten spp (Arg i1), Argas spp (Arg r 1, Arg r 1.0101), Ariopsis spp (Ari fe 1),Armoracia spp (Arm r HRP), Arrhenatherum spp (Arr e 1, Arr e 5),Artemisia spp (Art a 1, Art ap 1), Artemia spp (Art fr 1, Art fr 1.0101,Art fr 5, Art fr 5.0101), Arthrobacter spp (Art gl CO), Achorion spp(Art gy 7), Artocarpus spp (Art h 17kD, Art h 4), Arthrospira spp (ArtpI beta_Phycocyanin), Artemisia spp (Art v 1, Art v 1.0101, Art v1.0102, Art v 1.0103, Art v 1.0104, Art v 1.0105, Art v 1.0106, Art v1.0107, Art v 2, Art v 2.0101, Art v 3, Art v 3.0101, Art v 3.0201, Artv 3.0202, Art v 3.0301, Art v 4, Art v 4.0101, Art y 4.0201, Art v 47kD,Art v 5, Art v 5.0101, Art v 6, Art v 6.0101, Art v 60kD), Arthrodermaspp (Art va 4), Ascaris spp (Asc I 3, Asc I 3.0101, Asc I 3.0102, Asc I34kD, Asc s 1, Asc s 1.0101, Asc s 3, Asc s 3.0101, Asc s GST),Aspergillus spp (Asp aw Glucoamylase, Asp c 22, Asp f 1, Asp f 1.0101,Asp f 10, Asp f 10.0101, Asp f 11, Asp f 11.0101, Asp f 12, Asp f12.0101, Asp f 13, Asp f 13.0101, Asp f 15, Asp f 15.0101, Asp f 16, Aspf 16.0101, Asp f 17, Asp f 17.0101, Asp f 18, Asp f 18.0101, Asp f 2,Asp f 2.0101, Asp f 22, Asp f 22.0101, Asp f 23, Asp f 23.0101, Asp f27, Asp f 27.0101, Asp f 28, Asp f 28.0101, Asp f 29, Asp f 29.0101, Aspf 3, Asp f 3.0101, Asp f 34, Asp f 34.0101, Asp f 4, Asp f 4.0101, Asp f5, Asp f 5.0101, Asp f 56kD, Asp f 6, Asp f 6.0101, Asp f 7, Asp f7.0101, Asp f 8, Asp f 8.0101, Asp f 9, Asp f 9.0101, Asp f AfCalAp, Aspf AT_V, Asp f Catalase, Asp f Chitosanase, Asp f CP, Asp f DPPV, Asp fFDH, Asp f gamma_Actin, Asp f Glucosidase, Asp f GPI, Asp f GST, Asp fGT, Asp f IAO, Asp f IPMI, Asp f LPL1, Asp f LPL3, Asp f Mannosidase,Asp f MDH, Asp f PL, Asp f PUP, Asp f RPS3, Asp f SXR, Asp fl 13, Asp fl13.0101, Asp fl 18, Asp fl 2, Asp fl 21, Asp fl 3, Asp fl 4, Asp fl 7,Asp fl 8, Asp fl 9, Asp me Seaprose, Asp n 14, Asp n 14.0101, Asp n 18,Asp n 18.0101, Asp n 25, Asp n 25.0101, Asp n 30, Asp n Glucoamylase,Asp n Hemicellulase, Asp n Pectinase, Asp o 13, Asp o 13.0101, Asp o 21,Asp o 21.0101, Asp o 3, Asp o 4, Asp o 7, Asp o 8, Asp o Lactase, Asp oLipase, Asp oc 13, Asp r 1, Asp sa AP, Asp sp Glucoamylase, Asp spGlucoseoxidase, Asp sp PL, Asp sp PME, Asp sy 13, Asp v 13, Asp v13.0101, Asp v Catalase A, Asp v Enolase, Asp v GAPDH, Asp v MDH, Asp vSXR), Asparagus spp (Aspa o 1, Aspa o 1.01, Aspa o 1.02, Aspa o 17kD,Aspa o 4), Aspergillus spp (Aspe ni 2, Aspe ni 3, Aspe ni 4, Aspe ni 7,Aspe ni 8, Aspe ni 9), Avena spp (Ave s 1, Ave s 12, Ave s 13, Ave s 2,Ave s 4, Ave s 5, Ave s 7), Babylonia spp (Bab ja 1), Bacillus spp (Bacat Subtilisin, Bac cl Subtilisin, Bac I Subtilisin, Bac li aA, Bac liSubtilisin), Bactrocera spp (Bac of 27, Bac of 27.0101), Bacillus spp(Bac sp aA1, Bac sp aA3, Bac sp Decarboxylase, Bac st amyM, Bac suSubtilisin, Bac t Cryl Ab, Bac t Cryl Fa, Bac t Cry3Bbl, Bac t Cry9c),Bagre spp (Bag ma 1), Balistes spp (Bal ca 1), Balanus spp (Bal r 1, Balr 1.0101), Beauveria spp (Bea b Ald, Bea b Enol, Bea b f2, Bea b Hex),Bertholletia spp (Ber e 1, Ber e 1.0101, Ber e 2, Ber e 2.0101), Beryxspp (Ber sp 1), Betula spp (Bet ab 1, Bet al 1, Bet ch 1, Bet co 1, Betda 1, Bet gr 1, Bet hu 1, Bet le 1, Bet me 1, Bet n 1, Bet p 1, Bet pa1, Bet po 1, Bet pu 1, Bet pu 2, Bet pu 4, Bet pu 6, Bet pu 7, Bet sc 1,Bet ut 1, Bet v 1, Bet v 1B1-B1-B1, Bet v 1 fv Mal 4x, Bet v 1.0101, Betv 1.0102, Bet v 1.0103, Bet v 1.0201, Bet v 1.0301, Bet v 1.0401, Bet v1.0402, Bet v 1.0501, Bet v 1.0601, Bet v 1.0602, Bet v 1.0701, Bet v1.0801, Bet v 1.0901, Bet v 1.1001, Bet v 1.1101, Bet v 1.1201, Bet v1.1301, Bet v 1.1401, Bet v 1.1402, Bet v 1.1501, Bet v 1.1502, Bet v1.1601, Bet v 1.1701, Bet v 1.1801, Bet v 1.1901, Bet v 1.2001, Bet v1.2101, Bet v 1.2201, Bet v 1.2301, Bet v 1.2401, Bet v 1.2501, Bet v1.2601, Bet v 1.2701, Bet v 1.2801, Bet v 1.2901, Bet v 1.3001, Bet v1.3101, Bet v 2, Bet v 2.0101, Bet v 3, Bet v 3.0101, Bet v 4, Bet v4.0101, Bet v 6, Bet v 6.0101, Bet v 6.0102, Bet v 7, Bet v 7.0101, Betv 8, Bet v Glucanase), Beta spp (Beta v 1, Beta v 1.0101, Beta v 2, Betav 2.0101), Blattella spp (Bla g 1, Bla g 1.0101, Bla g 1.0102, Bla g1.0103, Bla g 1.0201, Bla g 1.0202, Bla g 2, Bla g 2.0101, Bla g 2.0201,Bla g 36kD, Bla g 4, Bla g 4.0101, Bla g 4.0201, Bla g 5, Bla g 5.0101,Bla g 5.0201, Bla g 6, Bla g 6.0101, Bla g 6.0201, Bla g 6.0301, Bla g7, Bla g 7.0101, Bla g 8, Bla g 8.0101, Bla g 9, Bla g Enolase, Bla gGSTD1, Bla g RACK1, Bla g TPI, Bla g Trypsin, Bla g Vitellogenin),Blatta spp (Bla o 1, Bla o 7), Blomia spp (Blo t 1, Blo t 1.0101, Blo t1.0201, Blo t 10, Blo t 10.0101, Blo t 10.0102, Blo t 11, Blo t 11.0101,Blo t 12, Blo t 12.0101, Blo t 12.0102, Blo t 13, Blo t 13.0101, Blo t14, Blo t 15, Blo t 18, Blo t 19, Blo t 19.0101, Blo t 2, Blo t 2.0101,Blo t 2.0102, Blo t 2.0103, Blo t 20, Blo t 21, Blo t 21.0101, Blo t 3,Blo t 3.0101, Blo t 4, Blo t 4.0101, Blo t 5, Blo t 5.0101, Blo t 6, Blot 6.0101, Blo t 7, Blo t 8, Blo t 9, Blo t HSP70), Bombus spp (Born ar4, Born by 4, Born p 1, Born p 1.0101, Born p 2, Born p 3, Born p 4,Born p 4.0101, Born t 1, Born t 1.0101, Born t 4, Born t 4.0101), Bombyxspp (Bomb m 1, Bomb m 1.0101, Bomb m 7, Bomb m 7.0101, Bomb m 7.0102,Bomb m 7.0103, Bomb m 7.0104, Bomb m 7.0105, Bomb m 7.0106), Boophilusspp (Boo m 1, Boo m 7, Boo m 7.0101), Bos spp (Bos d 2, Bos d 2.0101,Bos d 2.0102, Bos d 2.0103, Bos d 3, Bos d 3.0101, Bos d 4, Bos d4.0101, Bos d 5, Bos d 5.0101, Bos d 5.0102, Bos d 6, Bos d 6 (MDA), Bosd 6.0101, Bos d 7, Bos d 7.0101, Bos d 8, Bos d 8 alphaS1, Bos d 8alphaS2, Bos d 8 beta, Bos d 8 kappa, Bos d alpha2l, Bos d alpha2l.0101,Bos d Chymosin, Bos d Fibrin, Bos d Gelatin, Bos d HG, Bos d Insulin,Bos d Lactoferrin, Bos d Lactoperoxidase, Bos d Myoglobin, Bos d OBP,Bos d OSCP, Bos d Phosvitin, Bos d PLA2, Bos d PRVB, Bos d Thrombin, Bosd TI, Bos gr ALA, Bos gr Myoglobin), Bothrops spp (Bot as 1, Bot at 1),Bouteloua spp (Bou g 1), Biting spp (Boy ov 1), Brama spp (Bra du 1),Brassica spp (Bra j 1, Bra j 1.0101, Bra n 1, Bra n 1.0101, Bra n 4, Bran 7, Bra n 8, Bra n PG, Bra ni 8, Bra o 3, Bra o 3.0101, Bra r 1, Bra r1.0101, Bra r 2, Bra r 2.0101, Bra r 3, Bra r 4, Bra r 7), Bromus spp(Bro a 1, Bro a 4), Brosme spp (Bro br 1), Bromus spp (Bro i 1, Bro i 5,Bro i 7), Brugia spp (Bru m 3, Bru m 3.0101, Bru m Bm33), Bubalus spp(Bub b ALA, Bub b BLG, Bub b Casein, Bub b Casein alphaS1, Bub b CaseinalphaS2, Bub b Casein beta, Bub b Casein kappa), Caenorhabditis spp (Caeb 3, Cae b 3.0101, Cae br 3, Cae br 3.0101, Cae e 3, Cae e 3.0101, Cae e3.0102, Cae re 13, Cae re 13.0101), Cajanus spp (Caj c 1), Caligus spp(Cal cl 1, Cal cl 1.0101, Cal cl 1.0102), Calamus spp (Cal le 1),Callinectes spp (Cal s 2), Camelus spp (Cam d ALA, Cam d Casein, Cam dCasein alphaS1, Cam d Casein alphaS2, Cam d Casein beta, Cam d Caseinkappa), Camponotus spp (Cam fl 7, Cam fl 7.0101), Canis spp (Can f 1,Can f 1.0101, Can f 2, Can f 2.0101, Can f 3, Can f 3.0101, Can f 4, Canf 4.0101, Can f 5, Can f 5.0101, Can f 6, Can f 6.0101, Can fFeld1-like, Can f Homs2-like, Can f Phosvitin, Can f TCTP), Canthidermisspp (Can ma 1), Cancer spp (Can mg 2, Can p 1), Cannabis spp (Can s 3),Candida spp (Cand a 1, Cand a 1.0101, Cand a 3, Cand a 3.0101, Cand aCAAP, Cand a CyP, Cand a Enolase, Cand a FPA, Cand a MnSOD, Cand a PGK,Cand b 2, Cand b 2.0101, Cand b FDH, Cand r Lipase), Capsicum spp (Cap a1, Cap a 1.0101, Cap a 17kD, Cap a 2, Cap a 2.0101, Cap a 30kD, Cap aGlucanase, Cap ch 17kD), Caprella spp (Cap e 1), Capra spp (Cap h ALA,Cap h BLG, Cap h Casein, Cap h Casein alphaS1, Cap h Casein alphaS2, Caph Casein beta, Cap h Casein kappa, Cap h GSA), Capitulum spp (Cap m 1),Carassius spp (Car au 1), Carpinus spp (Car b 1, Car b 1.0101, Car b1.0102, Car b 1.0103, Car b 1.0104, Car b 1.0105, Car b 1.0106, Car b1.0107, Car b 1.0108, Car b 1.0109, Car b 1.0110, Car b 1.0111, Car b1.0112, Car b 1.0113, Car b 1.0201, Car b 1.0301, Car b 1.0302, Car b 2,Car b 4), Caranx spp (Car cr 1), Carya spp (Car i 1, Car i 1.0101, Car i2, Car i 4, Car i 4.0101), Carcinus spp (Car ma 2), Caryota spp (Car mi2), Carica spp (Car p 1, Car p Chitinase, Car p Chymopapain, Car pEndoproteinase), Castanea spp (Cas c 24kD, Cas s 1, Cas s 1.0101, Cas s1.0102, Cas s 1.0103, Cas s 2, Cas s 5, Cas s 5.0101, Cas s 8, Cas s8.0101, Cas s 9, Cas s 9.0101), Catharanthus spp (Cat r 1, Cat r 1.0101,Cat r 17kD, Cat r 2), Caulolatilus spp (Cau ch 1), Cavia spp (Cav p 1,Cav p 1.0101, Cav p 2, Cav p 2.0101, Cav p 3, Cav p 3.0101, Cav pGelatin, Cav p GSA), Centropristis spp (Cen s 1), Cephalopholis spp (Cepso 1), Charybdis spp (Cha f 1, Cha f 1.0101), Chaetodipterus spp (Cha fa1), Chamaecyparis spp (Cha o 1, Cha o 1.0101, Cha o 2, Cha o 2.0101),Chenopodium spp (Che a 1, Che a 1.0101, Che a 2, Che a 2.0101, Che a 3,Che a 3.0101), Chironomus spp (Chi k 1, Chi k 10, Chi k 10.0101),Chinchilla spp (Chi 121kD_a, Chi 121kD_b), Chionoecetes spp (Chi o 1,Chi o 1.0101, Chi o 2, Chi o 4, Chi o 6, Chi o alpha_Actin, Chi oSERCA), Chironomus spp (Chi t 1, Chi t 1.0101, Chi t 1.0201, Chi t 2,Chi t 2.0101, Chi t 2.0102, Chi t 3, Chi t 3.0101, Chi t 4, Chi t4.0101, Chi t 5, Chi t 5.0101, Chi t 6, Chi t 6.0101, Chi t 6.0201, Chit 7, Chi t 7.0101, Chi t 8, Chi t 8.0101, Chi t 9, Chi t 9.0101),Chlamys spp (Chl n 1), Chloephaga spp (Chl pi 1), Chortoglyphus spp (Choa 10), Chrysomela spp (Chr tr 7, Chr tr 7.0101), Cicer spp (Cic a 2SAlbumin, Cic a Albumin), Cichorium spp (Cic i 1), Cimex spp (Cim INitrophorin), Citrus spp (Cit I 1, Cit I 3, Cit I 3.0101), Citrullus spp(Cit la 2, Cit la MDH, Cit la TPI), Citrus spp (Cit r 3, Cit r 3.0101,Cit s 1, Cit s 1.0101, Cit s 2, Cit s 2.0101, Cit s 3, Cit s 3.0101, Cits 3.0102, Cit s IFR), Cladosporium spp (Cla c 14, CIa c 14.0101, Cla c9, CIa c 9.0101, Cla h 1, Cla h 10, Cla h 10.0101, Cla h 12, Cla h12.0101, Cla h 2, Cla h 2.0101, Cla h 42kD, Cla h 5, Cla h 5.0101, Cla h6, Cla h 6.0101, Cla h 7, Cla h 7.0101, Cla h 8, Cla h 8 CSP, Cla h8.0101, Cla h 9, Cla h 9.0101, Cla h abH, Cla h GST, Cla h HCh1, Cla hHSP70, Cla h NTF2, Cla h TCTP), Clostridium spp (Clo hi Collagenase, Clot Toxoid), Clupea spp (CIu h 1, Clu h 1.0101, CIu h 1.0201, CIu h1.0301), Cocos spp (Coc n 2, Coc n 4, Coc n 5), Coccidioides spp (Coc po8), Coffea spp (Cof a 1, Cof a 1.0101), Columba spp (Col I PSA),Coprinus spp (Cop c 1, Cop c 1.0101, Cop c 2, Cop c 2.0101, Cop c 3, Copc 3.0101, Cop c 4, Cop c 5, Cop c 5.0101, Cop c 6, Cop c 7, Cop c7.0101), Corylus spp (Cor a 1, Cor a 1.0101, Cor a 1.0102, Cor a 1.0103,Cor a 1.0104, Cor a 1.0201, Cor a 1.0301, Cor a 1.0401, Cor a 1.0402,Cor a 1.0403, Cor a 1.0404, Cor a 10, Cor a 10.0101, Cor a 11, Cor a11.0101, Cor a 12, Cor a 12.0101, Cor a 13, Cor a 13.0101, Cor a 14, Cora 14.0101, Cor a 2, Cor a 2.0101, Cor a 2.0102, Cor a 8, Cor a 8.0101,Cor a 9, Cor a 9.0101), Corynebacterium spp (Cor d Toxoid), Corylus spp(Cor he 1), Coryphaena spp (Cor hi 1), Coriandrum spp (Cor s 1, Cor s 11kD, Cor s 2), Cotoneaster spp (Cot I 3), Crangon spp (Cra c 1, Cra c1.0101, Cra c 2, Cra c 2.0101, Cra c 4, Cra c 4.0101, Cra c 5, Cra c5.0101, Cra c 6, Cra c 6.0101, Cra c 8, Cra c 8.0101), Crassostrea spp(Cra g 1), Cricetus spp (Cri c HSA), Crivellia spp (Cri pa 1), Crocusspp (Cro s 1, Cro s 1.0101, Cro s 2, Cro s 2.0101, Cro s 3, Cro s 3.01,Cro s 3.02), Cryptomeria spp (Cry j 1, Cry j 1.0101, Cry j 1.0102, Cry j1.0103, Cry j 2, Cry j 2.0101, Cry j 2.0102, Cry j 3, Cry j 3.1, Cry j3.2, Cry j 3.3, Cry j 3.4, Cry j 3.5, Cry j 3.6, Cry j 3.7, Cry j 3.8,Cry j 4, Cry j AP, Cry j Chitinase, Cry j CPA9, Cry j IFR, Cry j LTP,Cry j P1-P2), Cryphonectria spp (Cry p AP), Ctenocephalides spp (Cte f1, Cte f 1.0101, Cte f 2, Cte f 2.0101, Cte f 3, Cte f 3.0101),Ctenopharyngodon spp (Cte id 1), Cucumis spp (Cuc m 1, Cuc m 1.0101, Cucm 2, Cuc m 2.0101, Cuc m 3, Cuc m 3.0101, Cuc m Lec17, Cuc m MDH),Cucurbita spp (Cuc ma 18kD, Cuc ma 2, Cuc p 2, Cuc p AscO), Cucumis spp(Cuc s 2), Culicoides spp (Cul n 1, Cul n 10, Cul n 11, Cul n 2, Cul n3, Cul n 4, Cul n 5, Cul n 6, Cul n 7, Cul n 8, Cul n 9, Cul n HSP70),Culex spp (Cul q 28kD, Cul q 35kD, Cul q 7, Cul q 7.0101, Cul q 7.0102),Culicoides spp (Cul so 1), Cuminum spp (Cum c 1, Cum c 2), Cupressus spp(Cup a 1, Cup a 1.0101, Cup a 1.02, Cup a 2, Cup a 3, Cup a 4, Cup a4.0101, Cup s 1, Cup s 1.0101, Cup s 1.0102, Cup s 1.0103, Cup s 1.0104,Cup s 1.0105, Cup s 3, Cup s 3.0101, Cup s 3.0102, Cup s 3.0103, Cup s8), Cochliobolus spp (Cur 11, Cur I 1.0101, Cur 12, Cur I 2.0101, Cur13, Cur I 3.0101, Cur I 4, Cur I 4.0101, Cur I ADH, Cur I GST, Cur IMnSOD, Cur I Oryzin, Cur I Trx, Cur I ZPS1), Cyanochen spp (Cya cy 1),Cynoscion spp (Cyn ar 1), Cynosurus spp (Cyn cr 1, Cyn cr 5), Cynodonspp (Cyn d 1, Cyn d 1.0101, Cyn d 1.0102, Cyn d 1.0103, Cyn d 1.0104,Cyn d 1.0105, Cyn d 1.0106, Cyn d 1.0107, Cyn d 1.0201, Cyn d 1.0202,Cyn d 1.0203, Cyn d 1.0204, Cyn d 10, Cyn d 11, Cyn d 12, Cyn d 12.0101,Cyn d 13, Cyn d 15, Cyn d 15.0101, Cyn d 2, Cyn d 22, Cyn d 22.0101, Cynd 23, Cyn d 23.0101, Cyn d 24, Cyn d 24.0101, Cyn d 4, Cyn d 5, Cyn d 6,Cyn d 7, Cyn d 7.0101), Cynoscion spp (Cyn ne 1), Cynomys spp (Cyn spLipocalin), Cyprinus spp (Cyp c 1, Cyp c 1.01, Cyp c 1.02), Daboia spp(Dab ru 1), Dactylis spp (Dac g 1, Dac g 1.01, Dac g 1.0101, Dac g 1.02,Dac g 12, Dac g 13, Dac g 2, Dac g 2.0101, Dac g 3, Dac g 3.0101, Dac g4, Dac g 4.0101, Dac g 5, Dac g 5.0101, Dac g 7), Dama spp (Dam d CSA),Danio spp (Dan re 1, Dan re 2, Dan re alpha2l, Dan re CK), Dasyatis spp(Das ak 1, Das am 1, Das sa 1), Daucus spp (Dau c 1, Dau c 1.0101, Dau c1.0102, Dau c 1.0103, Dau c 1.0104, Dau c 1.0105, Dau c 1.0201, Dau c1.0301, Dau c 3, Dau c 4, Dau c 4.0101, Dau c CyP), Decapterus spp (Decru 1), Dendronephthya spp (Den n 1, Den n 1.0101), Dermatophagoides spp(Der f 1, Der f 1.0101, Der f 1.0102, Der f 1.0103, Der f 1.0104, Der f1.0105, Der f 1.0106, Der f 1.0107, Der f 1.0108, Der f 1.0109, Der f1.0110, Der f 10, Der f 10.0101, Der f 10.0102, Der f 11, Der f 11.0101,Der f 13, Der f 13.0101, Der f 14, Der f 14.0101, Der f 15, Der f15.0101, Der f 16, Der f 16.0101, Der f 17, Der f 17.0101, Der f 18, Derf 18.0101, Der f 2, Der f 2.0101, Der f 2.0102, Der f 2.0103, Der f2.0104, Der f 2.0105, Der f 2.0106, Der f 2.0107, Der f 2.0108, Der f2.0109, Der f 2.0110, Der f 2.0111, Der f 2.0112, Der f 2.0113, Der f2.0114, Der f 2.0115, Der f 2.0116, Der f 2.0117, Der f 20, Der f 21,Der f 22, Der f 22.0101, Der f 3, Der f 3.0101, Der f 4, Der f 5, Der f6, Der f 6.0101, Der f 7, Der f 7.0101, Der f 8, Der f 9, Der f HSP70),Dermanyssus spp (Der g 10, Der g 10.0101), Dermatophagoides spp (Der m1, Der m 1.0101, Der p 1, Der p 1.0101, Der p 1.0102, Der p 1.0103, Derp 1.0104, Der p 1.0105, Der p 1.0106, Der p 1.0107, Der p 1.0108, Der p1.0109, Der p 1.0110, Der p 1.0111, Der p 1.0112, Der p 1.0113, Der p1.0114, Der p 1.0115, Der p 1.0116, Der p 1.0117, Der p 1.0118, Der p1.0119, Der p 1.0120, Der p 1.0121, Der p 1.0122, Der p 1.0123, Der p1.0124, Der p 10, Der p 10.0101, Der p 10.0102, Der p 10.0103, Der p 11,Der p 11.0101, Der p 13, Der p 14, Der p 14.0101, Der p 15, Der p 18,Der p 2, Der p 2.0101, Der p 2.0102, Der p 2.0103, Der p 2.0104, Der p2.0105, Der p 2.0106, Der p 2.0107, Der p 2.0108, Der p 2.0109, Der p2.0110, Der p 2.0111, Der p 2.0112, Der p 2.0113, Der p 2.0114, Der p2.0115, Der p 20, Der p 20.0101, Der p 21, Der p 21.0101, Der p 23, Derp 23.0101, Der p 3, Der p 3.0101, Der p 4, Der p 4.0101, Der p 5, Der p5.0101, Der p 5.0102, Der p 6, Der p 6.0101, Der p 7, Der p 7.0101, Derp 8, Der p 8.0101, Der p 9, Der p 9.0101, Der p 9.0102, Der p P1-P2, Derp P2-P1, Der s 1, Der s 2, Der s 3), Dianthus spp (Dia c RIP),Dicranopteris spp (Dic I 2S Albumin), Diospyros spp (Dio k 17kD, Dio k4, Dio k IFR), Dioscorea spp (Dio p TSP), Diplodus spp (Dip ho 1),Distichlis spp (Dis s 1, Dis s 7), Ditrema spp (Dit to 1),Dolichovespula spp (Dol a 1, Dol a 2, Dol a 5, Dol a 5.0101), Dolichosspp (Dol b Agglutinin), Dolichovespula spp (Dol m 1, Dol m 1.0101, Dol m1.02, Dol m 2, Dol m 2.0101, Dol m 5, Dol m 5.0101, Dol m 5.02),Drosophila spp (Dro an 7, Dro an 7.0101, Dro er 7, Dro er 7.0101, Dro er7.0102, Dro gr 7, Dro gr 7.0101, Dro gr 7.0102, Dro m 7, Dro m 7.0101,Dro m 7.0102, Dro m 7.0103, Dro m 7.0104, Dro m 7.0105, Dro m 7.0106,Dro m 7.0107, Dro m 7.0108, Dro m 7.0109, Dro m 7.0110, Dro m 7.0111,Dro m 7.0112, Dro m 7.0113, Dro m 9, Dro m MnSOD, Dro mo 7, Dro mo7.0101, Dro pp 7, Dro pp 7.0101, Dro se 7, Dro se 7.0101, Dro si 7, Drosi 7.0101, Dro si 7.0102, Dro vi 7, Dro vi 7.0101, Dro wi 7, Dro wi7.0101, Dro y 7, Dro y 7.0101, Dro y 7.0102, Dro y 7.0103), Echium spp(Ech p Cytochrome C), Elaeis spp (Ela g 2, Ela g Bd31kD), Elops spp (Elosa 1), Embellisia spp (Emb a 1, Emb i 1, Emb nz 1, Emb t 1), Engraulisspp (Eng e 1), Enteroctopus spp (Ent d 1), Epinephelus spp (Epi bl 1,Epi co 1, Epi fl 1, Epi mc 1, Epi mo 1), Epicoccum spp (Epi p 1, Epi p1.0101, Epi p 12kD, Epi p GST), Epinephelus spp (Epi po 1, Epi un 1),Equisetum spp (Equ a 17kD), Equus spp (Equ as 4, Equ as DSA, Equ bu 4,Equ c 1, Equ c 1.0101, Equ c 2, Equ c 2.0101, Equ c 2.0102, Equ c 3, Equc 3.0101, Equ c 4, Equ c 4.0101, Equ c 5, Equ c 5.0101, Equ c ALA, Equ cBLG, Equ c Casein, Equ c Casein beta, Equ c Casein kappa, Equ c PRVB,Equ he 4, Equ z ZSA), Erimacrus spp (Eri i 1, Eri i 1.0101, Eri i1.0102), Eriocheir spp (Eri s 1, Eri s 1.0101, Eri s 2), Erwinia spp(Erw ch Asparaginase), Escherichia spp (Esc c Asparaginase, Esc c betaGAL), Esox spp (Eso I 1), Euphausia spp (Eup p 1, Eup p 1.0101),Euphasia spp (Eup s 1, Eup s 1.0101), Euroglyphus spp (Eur m 1, Eur m1.0101, Eur m 1.0102, Eur m 1.0103, Eur m 10, Eur m 14, Eur m 14.0101,Eur m 2, Eur m 2.0101, Eur m 2.0102, Eur m 3, Eur m 3.0101, Eur m 4, Eurm 4.0101), Evynnis spp (Evy j 1), Fagopyrum spp (Fag e 1, Fag e 1.0101,Fag e 10kD, Fag e 19kD, Fag e 2, Fag e 2.0101, Fag e TI), Fagus spp (Fags 1, Fag s 1.0101, Fag s 2, Fag s 4), Fagopyrum spp (Fag t 1, Fag t10kD, Fag t 2, Fag t 2.0101), Felis spp (Fel d 1, Fel d 1.0101, Fel d 2,Fel d 2.0101, Fel d 3, Fel d 3.0101, Fel d 4, Fel d 4.0101, Fel d 5, Feld 5.0101, Fel d 6, Fel d 6.0101, Fel d 7, Fel d 7.0101, Fel d 8, Fel d8.0101, Fel d IgG), Fenneropenaeus spp (Fen c 1, Fen c 2, Fen me 1, Fenme 1.0101), Festuca spp (Fes e 1, Fes e 13, Fes e 4, Fes e 5, Fes e 7,Fes p 1, Fes p 13, Fes p 4, Fes p 4.0101, Fes p 5, Fes r 1, Fes r 5),Ficus spp (Fic c 17kD, Fic c 4, Fic c Ficin), Foeniculum spp (Foe v 1,Foe v 2), Forsythia spp (For s 1), Forcipomyia spp (For t 1, For t1.0101, For t 2, For t 2.0101, For t 7, For t FPA, For t Myosin, For tTPI), Fragaria spp (Fra a 1, Fra a 1.0101, Fra a 3, Fra a 3.0101, Fra a3.0102, Fra a 3.0201, Fra a 3.0202, Fra a 3.0203, Fra a 3.0204, Fra a3.0301, Fra a 4, Fra a 4.0101, Fra c 1), Fraxinus spp (Fra e 1, Fra e1.0101, Fra e 1.0102, Fra e 1.0201, Fra e 12, Fra e 2, Fra e 3, Fra e9), Fragaria spp (Fra v 1), Fusarium spp (Fus c 1, Fus c 1.0101, Fus c2, Fus c 2.0101, Fus c 3, Fus s 1, Fus s 45kD, Fus sp Lipase), Gadus spp(Gad c 1, Gad c 1.0101, Gad c APDH, Gad m 1, Gad m 1.0101, Gad m 1.0102,Gad m 1.0201, Gad m 1.0202, Gad m 45kD, Gad m Gelatin, Gad ma 1), Gallusspp (Gal d 1, Gal d 1.0101, Gal d 2, Gal d 2.0101, Gal d 3, Gal d3.0101, Gal d 4, Gal d 4.0101, Gal d 5, Gal d 5.0101, Gal d 6, Gal d6.0101, Gal d Apo I, Gal d Apo VI, Gal d GPI, Gal d HG, Gal d IgY, Gal dL-PGDS, Gal d Ovomucin, Gal d Phosvitin, Gal d PRVB, Gal la 4), Galleriaspp (Gal m 18kD, Gal m 24kD), Gallus spp (Gal so 4), Gammarus spp (Gam sTM), Gelonium spp (Gel m RIP), Geothelphusa spp (Geo de 1), Glossina spp(Glo m 5, Glo m 5.0101, Glo m 7, Glo m 7.0101, Glo m 7.0102, Glo m7.0103), Glycine spp (Gly a Bd30K, Gly ar Bd30K, Gly ca Bd30K, Gly clBd30K, Gly cu Bd30K, Gly cy Bd30K), Glycyphagus spp (Gly d 10, Gly d10.0101, Gly d 13, Gly d 2, Gly d 2.0101, Gly d 2.0201, Gly d 2.03, Glyd 2/Lep d 2 L1, Gly d 2/Lep d 2 L2, Gly d 2/Lep d 2 L3, Gly d 2/Lep d 2L4, Gly d 2/Lep d 2 R1, Gly d 2/Lep d 2 R2, Gly d 2/Lep d 2 R3, Gly d2/Lep d 2 R4, Gly d 2/Lep d 2 R5, Gly d 20, Gly d 3, Gly d 5, Gly d5.01, Gly d 5.02, Gly d 7, Gly d 8), Glycine spp (Gly f Bd30K, Gly IBd30K, Gly m 1, Gly m 1.0101, Gly m 1.0102, Gly m 2, Gly m 2.0101, Gly m2S Albumin, Gly m 3, Gly m 3.0101, Gly m 3.0102, Gly m 39kD, Gly m 4,Gly m 4.0101, Gly m 5, Gly m 5.0101, Gly m 5.0201, Gly m 5.0301, Gly m5.0302, Gly m 50kD, Gly m 6, Gly m 6.0101, Gly m 6.0201, Gly m 6.0301,Gly m 6.0401, Gly m 6.0501, Gly m 68kD, Gly m Agglutinin, Gly m Bd28K,Gly m Bd30K, Gly m Bd60K, Gly m CPI, Gly m EAP, Gly m TI, Gly mi Bd30K,Gly s Bd30K, Gly t Bd30K, Gly to Bd30K), Gossypium spp (Gos h Vicilin),Haemophilus spp (Hae in P6), Haemaphysalis spp (Hae I 7, Hae I 7.0101,Hae q 7, Hae q 7.0101), Haliotis spp (Hal a 1, Hal d 1, Hal di 1, Hal diPM, Hal m 1, Hal m 1.0101, Hal r 1, Hal r 49kD, Hal ru 1), Harmonia spp(Har a 1, Har a 1.0101, Har a 2, Har a 2.0101), Harpegnathos spp (Har sa7, Har sa 7.0101, Har sa 7.0102), Helianthus spp (Hel a 1, Hel a 1.0101,Hel a 2, Hel a 2.0101, Hel a 2S Albumin, Hel a 3, Hel a 3.0101, Hel a4), Helix spp (Hel ap 1, Hel as 1, Hel as 1.0101), Heligmosomoides spp(Hel p 3, Hel p 3.0101), Helianthus spp (Hel to 1), Hemanthias spp (Hemle 1), Hemifusus spp (Hem t 1), Heterodera spp (Het g 3, Het g 3.0101),Hevea spp (Hey b 1, Hey b 1.0101, Hey b 10, Hey b 10.0101, Hey b10.0102, Hey b 10.0103, Hey b 11, Hey b 11.0101, Hey b 11.0102, Hey b12, Hey b 12.0101, Hey b 13, Hey b 13.0101, Hey b 14, Hey b 14.0101, Heyb 2, Hey b 2.0101, Hey b 3, Hey b 3.0101, Hey b 4, Hey b 4.0101, Hey b5, Hey b 5.0101, Hey b 6, Hey b 6.01, Hey b 6.02, Hey b 6.0202, Hey b6.03, Hey b 7, Hey b 7.01, Hey b 7.02, Hey b 7.D2, Hey b 7.S2, Hey b 8,Hey b 8.0101, Hey b 8.0102, Hey b 8.0201, Hey b 8.0202, Hey b 8.0203,Hey b 8.0204, Hey b 9, Hey b 9.0101, Hey b Citrate binding Protein, Heyb GAPDH, Hey b HSP80, Hey b IFR, Hey b Proteasome subunit, Hey bRotamase, Hey b SPI, Hey b Trx, Hey b UDPGP), Hexagrammos spp (Hex of1), Hippoglossus spp (Hip h 1), Hippoglossoides spp (Hip pl 1),Hippoglossus spp (Hip st 1), Hirudo spp (Hir me Hirudin), Holcus spp(Hol I 1, Hol I 1.0101, Hol I 1.0102, Hol I 2, Hol I 4, Hol I 5, Hol I5.0101, Hol I 5.0201), Holocnemus spp (Hol pl 9, Hol pI Hemocyanin),Homarus spp (Horn a 1, Horn a 1.0101, Horn a 1.0102, Horn a 1.0103, Horna 3, Horn a 3.0101, Horn a 4, Horn a 6, Horn a 6.0101, Horn g 1, Hom g2), Homo spp (Hom s 1, Hom s 1.0101, Hom s 2, Hom s 2.0101, Hom s 3, Homs 3.0101, Hom s 4, Hom s 4.0101, Hom s 5, Hom s 5.0101, Hom s AAT, Hom sACTH, Hom s Adalimumab, Hom s ALA, Hom s alpha_Actin, Hom salpha-Galactosidase, Hom s APDH, Hom s Arylsulfatase B, Hom s Casein,Hom s CyP A, Hom s CyP B, Hom s CyP C, Hom s DSF70, Hom s DSG3, Hom selF6, Hom s Etanercept, Hom s Factor IX, Hom s Factor VII, Hom s FactorVIII, Hom s G-CSF, Hom s Glucocerebrosidase, Hom s Glucosidase, Hom sHLA-DR-alpha, Hom s HSA, Hom s Iduronidase, Hom s Idursulfase, Hom sIgA, Hom s Insulin, Hom s Lactoferrin, Hom s Laminin gamma_(—)2, Hom sMnSOD, Hom s Oxytocin, Hom s P2, Hom s Phosvitin, Hom s Profilin, Hom sPSA, Hom s RP1, Hom s TCTP, Hom s TL, Hom s TPA, Hom s TPO, Hom sTransaldolase, Hom s Trx, Hom s Tubulin-alpha, Hom s/Mus m Basiliximab,Hom s/Mus m Cetuximab, Hom s/Mus m Cetuximab (Gal-Gal), Hom s/Mus mInfliximab, Hom s/Mus m Natalizumab, Hom s/Mus m Omalizumab, Hom s/Mus mPalivizumab, Hom s/Mus m Rituximab, Hom s/Mus m Tocilizumab, Hom s/Mus mTrastuzumab), Hoplostethus spp (Hop a 1), Hordeum spp (Hor v 1, Hor v12, Hor v 12.0101, Hor v 13, Hor v 14, Hor v 15, Hor v 15.0101, Hor v16, Hor v 16.0101, Hor v 17, Hor v 17.0101, Hor v 18kD, Hor v 2, Hor v21, Hor v 21.0101, Hor v 28, Hor v 33, Hor v 4, Hor v 5, Hor v 5.0101,Hor v BDAI, Hor v BTI), Humicola spp (Hum in Cellulase), Humulus spp(Hum j 1, Hum j 1.0101, Hum j 10kD, Hum j 2), Huso spp (Hus h 1),Hylocereus spp (Hyl un LTP), Hymenocephalus spp (Hym st 1), Hyperoglyphespp (Hyp by 1), Hypophthalmichthys spp (Hyp mo 1), Hypophthalmichthy spp(Hyp no 1), Ictalurus spp (Ict fu 1, Ict p 1), Imperata spp (Imp c 4,Imp c 5, Imp c Mel), Ixodes spp (Ixo r 2, Ixo sc 7, Ixo sc 7.0101),Jasus spp (Jas Ia 1, Jas Ia 1.0101, Jas Ia 1.0102), Juglans spp (Jug ca1, Jug ca 2, Jug ci 1, Jug ci 2, Jug n 1, Jug n 1.0101, Jug n 2, Jug n2.0101, Jug r 1, Jug r 1.0101, Jug r 2, Jug r 2.0101, Jug r 3, Jug r3.0101, Jug r 4, Jug r 4.0101, Jug r 5), Juniperus spp (Jun a 1, Jun a1.0101, Jun a 1.0102, Jun a 2, Jun a 2.0101, Jun a 3, Jun a 3.0101, Junc 1, Jun o 1, Jun o 4, Jun o 4.0101, Jun r 3, Jun r 3.1, Jun r 3.2, Junv 1, Jun v 1.0101, Jun v 1.0102, Jun v 3, Jun v 3.0101, Jun v 3.0102,Jun v 4), Katsuwonus spp (Kat p 1), Kyphosus spp (Kyp se 1),Lachnolaimus spp (Lac ma 1), Lachesis spp (Lac mu 1), Lactuca spp (Lac s1, Lac s 1.0101), Lagocephalus spp (Lag Ia 1), Larus spp (Lar a 1, Lar a2, Lar a 3), Larimichthys spp (Lar po 1), Lates spp (Lat c 1),Lateolabrax spp (Lat ja 1), Lathyrus spp (Lat oc Agglutinin), Leiostomusspp (Lei xa 1), Lens spp (Len c 1, Len c 1.0101, Len c 1.0102, Len c1.0103, Len c 2, Len c 2.0101, Len c 3, Len c 3.0101, Len c Agglutinin),Leopardus spp (Leo p 1), Lepidoglyphus spp (Lep d 10, Lep d 10.0101, Lepd 12, Lep d 13, Lep d 13.0101, Lep d 2, Lep d 2.0101, Lep d 2.0102, Lepd 2.0201, Lep d 2.0202, Lep d 3, Lep d 39kD, Lep d 5, Lep d 5.0101, Lepd 5.0102, Lep d 5.0103, Lep d 7, Lep d 7.0101, Lep d 8, Lep d alphaTubulin), Lepomis spp (Lep gi 1), Leptomelanosoma spp (Lep i 1), Lepomisspp (Lep ma 1), Lepisma spp (Lep s 1, Lep s 1.0101, Lep s 1.0102),Lepeophtheirus spp (Lep sa 1, Lep sa 1.0101, Lep sa 1.0102, Lep sa1.0103), Leptailurus spp (Lep se 1), Lepidorhombus spp (Lep w 1, Lep w1.0101), Lethocerus spp (Let in 7, Let in 7.0101, Let in 7.0102),Leuciscus spp (Leu ce 1), Lewia spp (Lew in 1), Ligustrum spp (Lig v 1,Lig v 1.0101, Lig v 1.0102, Lig v 2), Lilium spp (Lil I 2, Lil I PG),Limanda spp (Lim fe 1), Limnonectes spp (Lim m 1), Limulus spp (Lim p 1,Lim p 1.0101, Lim p 2, Lim p LPA), Liposcelis spp (Lip b 1, Lip b1.0101), Litchi spp (Lit c 1, Lit c 1.0101, Lit c IFR, Lit c TPI),Lithobates spp (Lit ca 1), Litopenaeus spp (Lit se 1, Lit v 1, Lit v1.0101, Lit v 2, Lit v 2.0101, Lit v 3, Lit v 3.0101, Lit v 4, Lit v4.0101), Filiaria spp (Loa lo 3, Loa lo 3.0101), Lobotes spp (Lob su 1),Locusta spp (Loc m 7, Loc m 7.0101), Loligo spp (Lol b 1, Lol e 1),Lolium spp (Lol m 2, Lol m 5, Lol p 1, Lot p 1.0101, Lol p 1.0102, Lol p1.0103, Lol p 10, Lol p 11, Lol p 11.0101, Lol p 12, Lol p 13, Lol p 2,Lol p 2.0101, Lot p 3, Lol p 3.0101, Lol p 4, Lol p 4.0101, Lot p 5, Lotp 5.0101, Lol p 5.0102, Lol p 7, Lol p CyP, Lol p FT, Lol p Legumin),Lonomia spp (Lon o 7, Lon o 7.0101), Lophodytes spp (Lop cu 1),Lophonetta spp (Lop sp 1), Lupinus spp (Lup a 1, Lup a atpha_Congtutin,Lup a delta_Conglutin, Lup a gamma_Congtutin, Lup an 1, Lup an 1.0101,Lup an alpha_Conglutin, Lup an delta_Conglutin, Lup an gamma_Congtutin,Lup I 17kD), Lutjanus spp (Lut a 1, Lut c 1, Lut cy 1, Lut gr 1, Lut gu1, Lut jo 1), Lutraria spp (Lut p 1), Lutjanus spp (Lut pu 1, Lut sy 1),Lycopersicon spp (Lyc e 1, Lyc e 1.0101, Lyc e 11S Globulin, Lyc e 2,Lyc e 2.0101, Lyc e 2.0102, Lyc e 3, Lyc e 3.0101, Lyc e 4, Lyc e4.0101, Lyc e ARP60S, Lyc e Chitinase, Lyc e Glucanase, Lyc ePeroxidase, Lyc e PG, Lyc e PME, Lyc e PR23, Lyc e Vicilin),Maconellicoccus spp (Mac h 7, Mac h 7.0101), Macruronus spp (Mac ma 1,Mac n 1), Maclura spp (Mac po 17kD), Macrobrachium spp (Mac ro 1, Mac ro1.0101, Mac ro Hemocyanin), Macropus spp (Macy s Gelatin), Malus spp(Mal d 1, Mal d 1.0101, Mal d 1.0102, Mal d 1.0103, Mal d 1.0104, Mal d1.0105, Mal d 1.0106, Mal d 1.0107, Mal d 1.0108, Mal d 1.0109, Mal d1.0201, Mal d 1.0202, Mal d 1.0203, Mal d 1.0204, Mal d 1.0205, Mal d1.0206, Mal d 1.0207, Mal d 1.0208, Mal d 1.0301, Mal d 1.0302, Mal d1.0303, Mal d 1.0304, Mal d 1.0401, Mal d 1.0402, Mal d 1.0403, Mal d 2,Mal d 2.0101, Mal d 3, Mal d 3.0101, Mal d 3.0102, Mal d 3.0201, Mal d3.0202, Mal d 3.0203, Mal d 4, Mal d 4.0101, Mal d 4.0102, Mal d 4.0201,Mal d 4.0202, Mal d 4.0301, Mal d 4.0302), Malpighia spp (Mal g 4, Mal gHevein), Malus spp (Mal p 1), Malassezia spp (Mala f 2, Mala f 2.0101,Mala f 3, Mala f 3.0101, Mala f 4, Mala f 4.0101, Mala g 10, Mala s 1,Mala s 1.0101, Mala s 10, Mala s 10.0101, Mala s 11, Mala s 11.0101,Mala s 12, Mala s 12.0101, Mala s 13, Mala s 13.0101, Mala s 5, Mala s5.0101, Mala s 6, Mala s 6.0101, Mala s 7, Mala s 7.0101, Mala s 8, Malas 8.0101, Mala s 9, Mala s 9.0101), Manihot spp (Man e 5, Man e 5.0101,Man e FPA, Man e GAPDH), Mangifera spp (Man i 1, Man i 14kD, Man i 2,Man i 3, Man i 3.01, Man i 3.02, Man i Chitinase), Marsupenaeus spp (Marj 1, Mar j 1.0101, Mar j 2, Mar j 4), Matricaria spp (Mat c 17kD),Mecopoda spp (Mec e 7), Megalobrama spp (Meg am 2, Meg am CK), Megathuraspp (Meg c Hemocyanin), Megalops spp (Meg sp 1), Melanogrammus spp (Mela 1), Meleagris spp (Mel g 1, Mel g 2, Mel g 3, Mel g PRVB, Mel g TSA),Melicertus spp (Mel 11), Menticirrhus spp (Men am 1), Mercurialis spp(Mer a 1, Mer a 1.0101), Merluccius spp (Mer ap 1, Mer au 1, Mer bi 1,Mer ca 1, Mer ga 1, Mer hu 1), Merlangius spp (Mer me 1), Merluccius spp(Mer mr 1, Mer pa 1, Mer po 1, Mer pr 1, Mer se 1), Meriones spp (Mer un23kD), Metarhizium spp (Met a 30), Metapenaeopsis spp (Met ba 1),Metapenaeus spp (Met e 1, Met e 1.0101, Met e 2), Metasequoia spp (Metgl 2), Metapenaeus spp (Met j 1, Met j 2), Metanephrops spp (Met ja 1),Metapenaeopsis spp (Met la 1), Metanephrops spp (Met t 2),Micromesistius spp (Mic po 1), Micropogonias spp (Mic un 1), Mimachlamysspp (Mim n 1), Momordica spp (Mom c RIP), Morus spp (Mor a 17kD, Mor a4), Morone spp (Mor am 1), Morus spp (Mor n 3, Mor n 3.0101), Morone spp(Mor sa 1, Mor sc 1), Mugil spp (Mug c 1), Muraenolepis spp (Mur mi 1),Musa spp (Mus a 1, Mus a 1.0101, Mus a 2, Mus a 2.0101, Mus a 3, Mus a3.0101, Mus a 4, Mus a 4.0101, Mus a 5, Mus a 5.0101, Mus a 5.0102), Musspp (Mus m 1, Mus m 1.0101, Mus m 1.0102, Mus m 2, Mus m Gelatin, Mus mIgG, Mus m MSA, Mus m Muromonab, Mus m Phosvitin), Mustela spp (Mus p17kD), Musa spp (Mus xp 1, Mus xp 2, Mus xp 5), Mycteroperca spp (Myc bo1, Myc mi 1, Myc ph 1), Myceliophthora spp (Myc sp Laccase), Myrmeciaspp (Myr p 1, Myr p 1.0101, Myr p 2, Myr p 2.0101, Myr p 2.0102, Myr p3, Myr p 3.0101), Mytilus spp (Myt e 1, Myt g 1, Myt g PM), Myzus spp(Myz p 7, Myz p 7.0101), Nemorhedus spp (Nae go Hya), Necator spp (Nec aCalreticulin), Nemipterus spp (Nem vi 1), Neosartorya spp (Neo fi 1, Neofi 22), Neochen spp (Neo ju 1), Neoscona spp (Neo n 7, Neo n 7.0101),Nephelium spp (Nep I GAPDH), Nephrops spp (Nep n 1, Nep n DF9), Neptuneaspp (Nep po 1, Nep po 1.0101), Nicotiana spp (Nic t 8, Nic t Osmotin,Nic t Villin), Nimbya spp (Nim c 1, Nim s 1), Nippostrongylus spp (Nip bAg1), Nycticebus spp (Nyc c 1), Octopus spp (Oct f 1, Oct 11, Oct v 1,Oct v 1.0101, Oct v PM), Ocyurus spp (Ocy ch 1), Olea spp (Ole e 1, Olee 1.0101, Ole e 1.0102, Ole e 1.0103, Ole e 1.0104, Ole e 1.0105, Ole e1.0106, Ole e 1.0107, Ole e 10, Ole e 10.0101, Ole e 11, Ole e 11.0101,Ole e 11.0102, Ole e 12, Ole e 13, Ole e 2, Ole e 2.0101, Ole e 3, Ole e3.0101, Ole e 36kD, Ole e 4, Ole e 4.0101, Ole e 5, Ole e 5.0101, Ole e6, Ole e 6.0101, Ole e 7, Ole e 7.0101, Ole e 8, Ole e 8.0101, Ole e 9,Ole e 9.0101), Ommastrephes spp (Omm b 1, Omm b 1.0101), Oncorhynchusspp (Onc ke 1, Onc ke 18 kD, Onc ke alpha2l, Onc ke Vitellogenin, Onc m1, Onc m 1.0101, Onc m 1.0201, Onc m alpha2l, Onc m Protamine, Onc mVitellogenin, Onc ma 1, Onc ma FPA, Onc ma FSA, Onc ma TPI, Onc n 1),Onchocerca spp (Onc o 3, Onc o 3.0101), Oncorhynchus spp (Onc is 1),Onchocerca spp (Onc v 3, Onc v 3.0101), Oratosquilla spp (Ora o 1, Ora o1.0101), Oreochromis spp (Ore a 1, Ore mo 1, Ore mo 2, Ore mo FPA, Oremo SCAF7145, Ore ni 1, Ore ni 18kD, Ore ni 45kD), Ornithonyssus spp (Ornsy 10, Orn sy 10.0101, Orn sy 10.0102), Oryctolagus spp (Ory c 1, Ory c1.0101, Ory c 2, Ory c Casein, Ory c Phosvitin, Ory c RSA), Oryza spp(Ory s 1, Ory s 1.0101, Ory s 11, Ory s 12, Ory s 12.0101, Ory s 13, Orys 14, Ory s 17kD, Ory s 19kD, Ory s 2, Ory s 23, Ory s 3, Ory s 7, Ory saA_TI, Ory s GLP52, Ory s GLP63, Ory s Glyoxalase I, Ory s NRA), Ostryaspp (Ost c 1, Ost c 1.0101), Ovis spp (Ovi a ALA, Ovi a BLG, Ovi aCasein, Ovi a Casein alphaS1, Ovi a Casein alphaS2, Ovi a Casein beta,Ovi a Casein kappa, Ovi a Phosvitin, Ovi a SSA), Pachycondyla spp (Pac c3), Pagrus spp (Pag m 1, Pag pa 1), Pampus spp (Pam ar 1, Pam c 1),Pandalus spp (Pan b 1, Pan b 1.0101), Pangasius spp (Pan bo 1), Pandalusspp (Pan e 1, Pan e 1.0101, Pan e 4), Panulirus spp (Pan h 1, Pan hy 1),Pangasius spp (Pan hy 18kD, Pan hy 45kD), Panulirus spp (Pan j 1),Panthera spp (Pan 11, Pan o 1, Pan p 1), Panulirus spp (Pan s 1, Pan s1.0101), Panthera spp (Pan t 1), Pan spp (Pan tr TCTP), Papaver spp (Paps 17kD, Pap s 2, Pap s 34kD), Papilio spp (Pap xu 7, Pap xu 7.0101, Papxu 7.0102), Paralichthys spp (Par a 1), Parasilurus spp (Par as 1, Par c1), Paralithodes spp (Par c 1.0101, Par c 1.0102, Par f 1), Partheniumspp (Par h 1), Parietaria spp (Par j 1, Par j 1.0101, Par j 1.0102, Parj 1.0103, Par j 1.0201, Par j 2, Par j 2.0101, Par j 2.0102, Par j 3,Par j 3.0101, Par j 3.0102, Par j 4, Par j 4.0101, Par j J1-J2),Paralichthys spp (Par le 1), Parietaria spp (Par m 1, Par o 1, Par o1.0101), Paralichthys spp (Par of 1, Par of alpha21), Parahucho spp (Parpe Vitellogenin), Passiflora spp (Pas e Chitinase, Pas e Hevein),Paspalum spp (Pas n 1, Pas n 1.0101, Pas n 13), Patinopecten spp (Pat y1), Pediculus spp (Ped h 7, Ped h 7.0101), Penaeus spp (Pen a 1, Pen a1.0101, Pen a 1.0102, Pen a 1.0102 (103-117), Pen a 1.0102 (109-123),Pen a 1.0102 (1-15), Pen a 1.0102 (115-129), Pen a 1.0102 (121-135), Pena 1.0102 (127-141), Pen a 1.0102 (13-27), Pen a 1.0102 (133-147), Pen a1.0102 (139-153), Pen a 1.0102 (145-159)), Farfantepenaeus spp (Pen a1.0102 (151-165)), Penaeus spp (Pen a 1.0102 (157-171), Pen a 1.0102(163-177), Pen a 1.0102 (169-183), Pen a 1.0102 (175-189), Pen a 1.0102(181-195), Pen a 1.0102 (187-201), Pen a 1.0102 (193-207), Pen a 1.0102(19-33), Pen a 1.0102 (199-213), Pen a 1.0102 (205-219), Pen a 1.0102(211-225), Pen a 1.0102 (217-231), Pen a 1.0102 (223-237), Pen a 1.0102(229-243)), Farfantepenaeus spp (Pen a 1.0102 (235-249)), Penaeus spp(Pen a 1.0102 (241-255), Pen a 1.0102 (247-261), Pen a 1.0102 (253-267),Pen a 1.0102 (25-39), Pen a 1.0102 (259-273), Pen a 1.0102 (265-279),Pen a 1.0102 (270-284), Pen a 1.0102 (31-45), Pen a 1.0102 (37-51), Pena 1.0102 (43-57), Pen a 1.0102 (49-63)), Farfantepenaeus spp (Pen a1.0102 (55-69)), Penaeus spp (Pen a 1.0102 (61-75), Pen a 1.0102(67-81), Pen a 1.0102 (7-21), Pen a 1.0102 (73-87), Pen a 1.0102(79-93), Pen a 1.0102 (85-99), Pen a 1.0102 (91-105), Pen a 1.0102(97-111), Pen a 1.0103), Penicillium spp (Pen b 13, Pen b 13.0101, Pen b26, Pen b 26.0101, Pen c 1, Pen c 13, Pen c 13.0101, Pen c 18, Pen c 19,Pen c 19.0101, Pen c 2, Pen c 22, Pen c 22.0101, Pen c 24, Pen c24.0101, Pen c 3, Pen c 3.0101, Pen c 30, Pen c 30.0101, Pen c 32, Pen c32.0101, Pen c MnSOD, Pen ch 13, Pen ch 13.0101, Pen ch 18, Pen ch18.0101, Pen ch 20, Pen ch 20.0101, Pen ch 31, Pen ch 31.0101, Pen ch33, Pen ch 33.0101, Pen ch 35, Pen ch 35.0101, Pen ch MnSOD), Penaeusspp (Pen i 1, Pen i 1.0101, Pen m 1, Pen m 1.0101, Pen m 1.0102, Pen m2, Pen m 2.0101, Pen m 3, Pen m 3.0101, Pen m 4, Pen m 4.0101, Pen m 6,Pen m 6.0101), Penicillium spp (Pen o 18, Pen o 18.0101), Penaeus spp(Pena o 1, Pena o 1.0101), Periplaneta spp (Per a 1, Per a 1.0101, Per a1.0102, Per a 1.0103, Per a 1.0104, Per a 1.0105, Per a 1.0201, Per a10, Per a 10.0101, Per a 2, Per a 3, Per a 3.0101, Per a 3.0201, Per a3.0202, Per a 3.0203, Per a 4, Per a 5, Per a 6, Per a 6.0101, Per a 7,Per a 7.0101, Per a 7.0102, Per a 7.0103, Per a 9, Per a 9.0101, Per aCathepsin, Per a FABP, Per a Trypsin, Per f 1, Per f 7, Per f 7.0101),Perna spp (Per v 1), Persea spp (Pers a 1, Pers a 1.0101, Pers a 4),Petroselinum spp (Pet c 1, Pet c 2, Pet c 3), Phalaris spp (Pha a 1, Phaa 1.0101, Pha a 5, Pha a 5.0101, Pha a 5.02, Pha a 5.03, Pha a 5.04),Phaseolus spp (Pha v 3, Pha v 3.0101, Pha v 3.0201, Pha v aAI, Pha vaA1.0101, Pha v Chitinase, Pha v PHA, Pha v Phaseolin), Phleum spp (Phlp 1, Phl p 1.0101, Phl p 1.0102, Phl p 11, Phl p 11.0101, Phl p 12, Phlp 12.0101, Phl p 12.0102, Phl p 12.0103, Phl p 13, Phi p 13.0101, Phi p2, Phi p 2.0101, Phi p 3, Phi p 3.0101, Phi p 3.0102, Phl p 4, Phl p4.0101, Phl p 4.0102, Phl p 4.0201, Phl p 4.0202, Phl p 4.0203, Phl p4.0204, Phl p 5, Phl p 5.0101, Phl p 5.0102, Phl p 5.0103, Phl p 5.0104,Phl p 5.0105, Phl p 5.0106, Phl p 5.0107, Phl p 5.0108, Phl p 5.0109,Phl p 5.0201, Phl p 5.0202, Phl p 5.0203, Phl p 5.0204, Phl p 5.0205,Phl p 5.0206, Phl p 5.0207, Phl p 6, Phl p 6.0101, Phl p 6.0102, Phl p7, Phl p 7.0101, Phl p P1-P2-P5-P6, Phl p P2-P6, Phl p P5-P1, Phl pP6-P2), Phoenix spp (Pho d 2, Pho d 2.0101, Pho d 40kD, Pho d 90kD),Phodopus spp (Pho s 21kD), Phoma spp (Pho t 1), Phragmites spp (Phr a 1,Phr a 12, Phr a 13, Phr a 4, Phr a 5), Phytolacca spp (Phy a RIP),Pimpinella spp (Pim a 1, Pim a 2), Pinna spp (Pin a 1), Piper spp (Pip n14kD, Pip n 28kD), Pisum spp (Pis s 1, Pis s 1.0101, Pis s 1.0102, Pis s2, Pis s 2.0101, Pis s 5, Pis s Agglutinin, Pis s Albumin), Pistacia spp(Pis v 1, Pis v 1.0101, Pis v 2, Pis v 2.0101, Pis v 2.0201, Pis v 3,Pis v 3.0101, Pis v 4, Pis v 4.0101, Pis v 5, Pis v 5.0101), Platanusspp (Pla a 1, Pla a 1.0101, Pla a 2, Pla a 2.0101, PIa a 3, Pla a3.0101, Pla a 8), Platichthys spp (Pla f 1), Plantago spp (Pla I 1, PlaI 1.0101, Pla I 1.0102, Pla I 1.0103, Pla I Cytochrome C), Platanus spp(Pla oc 1, Pla or 1, PIa or 1.0101, Pla or 2, Pla or 2.0101, Pla or 3,Pla or 3.0101, Pla or 4, Pla or CyP, Pla r 1), Plectropomus spp (Ple ar1), Pleospora spp (Ple h 1), Plectropomus spp (Ple le 1), Plodia spp(Plo i 1, Plo i 1.0101, Plo i 2, Plo i 2.0101), Poa spp (Poa p 1, Poa p1.0101, Poa p 10, Poa p 12, Poa p 13, Poa p 2, Poa p 4, Poa p 5, Poa p5.0101, Poa p 6, Poa p 7), Polistes spp (Pol a 1, Pol a 1.0101, Pol a 2,Pol a 2.0101, Pol a 5, Pot a 5.0101, Pol d 1, Pot d 1.0101, Pot d1.0102, Pot d 1.0103, Pot d 1.0104, Pol d 4, Pol d 4.0101, Pol d 5, Pold 5.0101, Pol e 1, Pol e 1.0101, Pol e 2, Pol e 4, Pol e 4.0101, Pol e5, Pol e 5.0101, Pol f 5, Pol f 5.0101, Pol g 1, Pol g 1.0101, Pol g 2,Pol g 4, Pol g 5, Pol g 5.0101, Pol he MLT, Pol m 5, Pol m 5.0101),Polypedilum spp (Pol n 1), Pollicipes spp (Pol po 1), Pollachius spp(Pol vi 1), Polybia spp (Poly p 1, Poly p 1.0101, Poly p 2, Poly p 5,Poly s 5, Poly s 5.0101), Pomatomus spp (Porn sa 1), Pongo spp (Pon abHSA), Pontastacus spp (Pon I 4, Pon I 4.0101, Pon 17, Pon I 7.0101),Portunus spp (Por s 1, Por s 1.0101, Por s 1.0102, Por tr 1, Por tr1.0101), Protortonia spp (Pro ca 38kD), Procumbarus spp (Pro cl 1, Procl 1.0101, Pro cl 21kD), Prosopis spp (Pro j 20kD), Prunus spp (Pru ar1, Pru ar 1.0101, Pru ar 3, Pru ar 3.0101, Pru av 1, Pru av 1.0101, Pruav 1.0201, Pru av 1.0202, Pru av 1.0203, Pru av 2, Pru av 2.0101, Pru av3, Pru av 3.0101, Pru av 4, Pru av 4.0101, Pru c 1, Pru d 1, Pru d 2,Pru d 3, Pru d 3.0101, Pru d 4, Pru du 1, Pru du 2, Pru du 2S Albumin,Pru du 3, Pru du 3.0101, Pru du 4, Pru du 4.0101, Pru du 4.0102, Pru du5, Pru du 5.0101, Pru du 6, Pru du 6.0101, Pru du 6.0201, Pru duConglutin, Pru p 1, Pru p 1.0101, Pru p 2, Pru p 2.0101, Pru p 2.0201,Pru p 2.0301, Pru p 3, Pru p 3.0101, Pru p 3.0102, Pru p 4, Pru p4.0101, Pru p 4.0201, Pru sa 3), Psilocybe spp (Psi c 1, Psi c 1.0101,Psi c 2, Psi c 2.0101), Psoroptes spp (Pso o 1, Pso o 10, Pso o 10.0101,Pso o 11, Pso o 13, Pso o 14, Pso o 2, Pso o 21, Pso o 3, Pso o 5, Pso o7), Puma spp (Pum c 1), Punica spp (Pun g 3), Pyrus spp (Pyr c 1, Pyr c1.0101, Pyr c 3, Pyr c 3.0101, Pyr c 4, Pyr c 4.0101, Pyr c 5, Pyr c5.0101, Pyr py 2), Quercus spp (Que a 1, Que a 1.0101, Que a 1.0201, Quea 1.0301, Que a 1.0401, Que a 2, Que a 4), Rachycentron spp (Rac ca 1),Rana spp (Ran e 1, Ran e 1.0101, Ran e 2, Ran e 2.0101), Ranina spp (Ranra 1), Rangifer spp (Ran t BLG), Rattus spp (Rat n 1, Rat n 1.0101, Ratn Casein, Rat n Gelatin, Rat n IgG, Rat n Phosvitin, Rat n RSA, Rat nTransferrin), Rhizomucor spp (Rhi m AP), Rhizopus spp (Rhi nv Lipase,Rhi o Lipase), Rhomboplites spp (Rho au 1), Rhodotorula spp (Rho m 1,Rho m 1.0101, Rho m 2, Rho m 2.0101), Ricinus spp (Ric c 1, Ric c1.0101, Ric c 2, Ric c 3, Ric c 8, Ric c RIP), Rivulus spp (Riv ma 1),Robinia spp (Rob p 2, Rob p 4, Rob p Glucanase), Rosa spp (Ros r 3),Roystonea spp (Roy e 2), Rubus spp (Rub i 1, Rub i 1.0101, Rub i 3, Rubi 3.0101, Rub i Chitinase, Rub i CyP), Saccharomyces spp (Sac cCarboxypeptidase Y, Sac c CyP, Sac c Enolase, Sac c Glucosidase, Sac cInvertase, Sac c MnSOD, Sac c P2, Sac c Profilin), Salvelinus spp (Sal f1), Salsola spp (Sal k 1, Sal k 1.0101, Sal k 1.0201, Sal k 1.0301, Salk 1.0302, Sal k 2, Sal k 2.0101, Sal k 3, Sal k 3.0101, Sal k 4, Sal k4.0101, Sal k 4.0201, Sal k 5, Sal k 5.0101), Salvelinus spp (Sal leVitellogenin), Salmo spp (Sal s 1, Sal s 1.0101, Sal s 1.0201, Sal s 2,Sal s 2.0101, Sal s Gelatin), Sambucus spp (Sam n 1), Sander spp (San lu1), Saponaria spp (Sap o RIP), Sardinops spp (Sar m 1), Sarkidiornis spp(Sar ml 1), Sardina spp (Sar p 1), Sarcoptes spp (Sar s 1, Sar s 14, Sars 3, Sar s GST, Sar s PM), Sardinops spp (Sar sa 1, Sar sa 1.0101),Schistosoma spp (Sch j GST, Sch j PM, Sch j Sj22, Sch j Sj67, Sch maSm20, Sch ma Sm21, Sch ma Sm22, Sch ma Sm31), Sciaenops spp (Sci oc 1),Scomber spp (Sco a 1), Scombermorus spp (Sco ca 1), Scomberomorus spp(Sco g 1), Scomber spp (Sco j 1, Sco ma 1, Sco s 1), Scolopendra spp(Sco y 7, Sco y 7.0101), Scylla spp (Scy o 1, Scy o 1.0101, Scy o 2, Scypa 1, Scy pa 2, Scy s 1, Scy s 1.0101, Scy s 2), Sebastes spp (Seb fa 1,Seb in 1, Seb m 1, Seb m 1.0101, Seb m 1.0201), Secale spp (Sec c 1, Secc 12, Sec c 13, Sec c 2, Sec c 20, Sec c 20.0101, Sec c 20.0201, Sec c28, Sec c 3, Sec c 4, Sec c 4.0101, Sec c 4.0201, Sec c 5, Sec c 5.0101,Sec c aA_TI, Sec c aA_TI.0101), Senecio spp (Sen j MDH, Sen j PL), Sepiaspp (Sep e 1, Sep e 1.0101), Sepioteuthis spp (Sep l 1, Sep l 1.0101),Sepia spp (Sep m 1), Seriola spp (Ser d 1, Ser la 1), Sergestes spp (Serlu 1), Seriola spp (Ser q 1, Ser ri 1), Sesamum spp (Ses i 1, Ses i1.0101, Ses i 2, Ses i 2.0101, Ses i 3, Ses i 3.0101, Ses i 4, Ses i4.0101, Ses i 5, Ses i 5.0101, Ses i 6, Ses i 6.0101, Ses i 7, Ses i7.0101, Ses i 8), Shigella spp (Shi bo GST, Shi dy GST), Simulia spp(Sim vi 1, Sim vi 2, Sim vi 3, Sim vi 4, Sim vi 70kD), Sinapis spp (Sina 1, Sin a 1.0101, Sin a 1.0104, Sin a 1.0105, Sin a 1.0106, Sin a1.0107, Sin a 1.0108, Sin a 2, Sin a 2.0101, Sin a 3, Sin a 3.0101, Sina 4, Sin a 4.0101), Sinonovacula spp (Sin c 1, Sin c 1.0101), Solenopsisspp (Sol g 2, Sol g 2.0101, Sol g 3, Sol g 3.0101, Sol g 4, Sol g4.0101, Sol g 4.0201, Sol i 1, Sol i 1.0101, Sol i 2, Sol i 2.0101, Soli 3, Sol i 3.0101, Sol i 4, Sol i 4.0101), Solenocera spp (Sol me 1),Solenopsis spp (Sol r 1, Sol r 2, Sol r 2.0101, Sol r 3, Sol r 3.0101,Sol s 2, Sol s 2.0101, Sol s 3, Sol s 3.0101, Sol s 4), Solea spp (Solso 1, Sol so TPI), Solanum spp (Sola t 1, Sola t 1.0101, Sola t 2, Solat 2.0101, Sola t 3, Sola t 3.0101, Sola t 3.0102, Sola t 4, Sola t4.0101, Sola t 8, Sola t Glucanase), Sorghum spp (Sor b 1, Sor h 1, Sorh 1.0101, Sor h 12, Sor h 7), Sparus spp (Spa a 1), Sphyrna spp (Sph ti1), Spirulina spp (Spi mx beta_Phycocyanin), Spinacia spp (Spi o 2, Spio RuBisCO), Squilla spp (Squ ac 1, Squ ac 1.0101, Squ o 1, Squ o1.0101), Staphylococcus spp (Sta a FBP, Sta a SEA, Sta a SEB, Sta a SEC,Sta a SED, Sta a SEE, Sta a TSST), Stachybotrys spp (Sta c 3, Sta c3.0101, Sta c Cellulase, Sta c Hemolysin, Sta c SchS34, Sta c StachyraseA), Stemphylium spp (Ste b 1, Ste c 1, Ste v 1), Stolephorus spp (Sto i1), Struthio spp (Str c 1, Str c 2, Str c 3), Streptococcus spp (Str dyStreptokinase), Streptomyces spp (Str g Pronase), Streptococcus spp (Strpn PspC), Strongylocentrotus spp (Str pu 18kD, Str pu Vitellogenin),Streptococcus spp (Str py SPEA, Str py SPEC, Str py Streptokinase),Strongyloides spp (Str st 45kD), Streptomyces spp (Str v PAT), Styelaspp (Sty p 1), Suidasia spp (Sui m 1, Sui m 13, Sui m 2, Sui m 3, Sui m5, Sui m 5.01, Sui m 5.02, Sui m 5.03, Sui m 6, Sui m 7, Sui m 8, Sui m9), Sus spp (Sus s ACTH, Sus s ALA, Sus s Amylase, Sus s BLG, Sus sCasein, Sus s Casein alphaS1, Sus s Casein alphaS2, Sus s Casein beta,Sus s Casein kappa, Sus s Gelatin, Sus s HG, Sus s Insulin, Sus sLipase, Sus s Pepsin, Sus s Phosvitin, Sus s PRVB, Sus s PSA, Sus sTCTP), Syntelopodeuma spp (Syn y 7, Syn y 7.0101), Syringa spp (Syr v 1,Syr v 1.0101, Syr v 1.0102, Syr v 1.0103, Syr v 2, Syr v 3, Syr v3.0101), Tabanus spp (Tab y 1, Tab y 1.0101, Tab y 2, Tab y 2.0101, Taby 5, Tab y 5.0101), Tadorna spp (Tad ra 1), Talaromyces spp (Tal st 22,Tal st 3, Tal st 8), Taraxacum spp (Tar o 18kD), Taxodium spp (Tax d 2),Tegenaria spp (Teg d Hemocyanin), Teladorsagia spp (Tel ci 3),Thaumetopoea spp (Tha p 1, Tha p 1.0101, Tha p 2, Tha p 2.0101),Theragra spp (The c 1), Thermomyces spp (The I Lipase, The sp Lipase,The sp Xylanase), Thunnus spp (Thu a 1, Thu a 1.0101, Thu a Collagen,Thu al 1, Thu at 1, Thu o 1, Thu o Collagen), Thuja spp (Thu oc 3, Thu p1), Thunnus spp (Thu t 1, Thu to 1), Thyrsites spp (Thy at 1),Thyrophygus spp (Thy y 7, Thy y 7.0101), Todarodes spp (Tod p 1, Tod p1.0101, Tod p 1.0102), Toxoptera spp (Tox c 7, Tox c 7.0101), Toxocaraspp (Tox ca TES120, Tox ca TES26, Tox ca TES30), Toxoplasma spp (Tox gHSP70), Trachypenaeus spp (Tra c 1), Trachinotus spp (Tra ca 1),Trachurus spp (Tra j 1, Tra j Gelatin, Tra tr Gelatin), Triticum spp(Tri a 1, Tri a 10kD, Tri a 12, Tri a 12.0101, Tri a 12.0102, Tri a12.0103, Tri a 12.0104, Tri a 13, Tri a 14, Tri a 14.0101, Tri a14.0201, Tri a 15, Tri a 15.0101, Tri a 18, Tri a 18.0101, Tri a 19, Tria 19.0101, Tri a 2, In a 21, Tri a 21.0101, Tri a 23kd, Tri a 25, Tri a25.0101, Tri a 26, Tri a 26.0101, Tri a 27, Tri a 27.0101, Tri a 28, Tria 28.0101, Tri a 29, Tri a 29.0101, Tri a 29.0201, Tri a 3, Tri a 30,Tri a 30.0101, Tri a 31, Tri a 31.0101, Tri a 32, Tri a 32.0101, Tri a33, Tri a 33.0101, Tri a 34, Tri a 34.0101, Tri a 35, Tri a 35.0101, Tria 36, Tri a 36.0101, Tri a 37, Tri a 37.0101, Tri a 4, Tri a 4.0101, Tria 4.0201, Tri a 5, Tri a 7, Tri a aA_SI, Tri a alpha_Gliadin, Tri a bA,Tri a Bd36K, Tri a beta_Gliadin, Tri a Chitinase, Tri a CM16, Tri a DH,Tri a Endochitinase, Tri a gamma_Gliadin, Tri a Germin, Tri a Gliadin,Tri a GST, Tri a LMW Glu, Tri a LMW-GS B16, Tri a LMW-GS P42, Tri aLMW-GS P73, Tri a LTP2, Tri a omega2_Gliadin, Tri a Peroxidase, Tri aPeroxidase 1, Tri a SPI, Tri a TLP, Tri a Tritin, Tri a XI),Tritirachium spp (Tri al Proteinase K), Tribolium spp (Tri ca 17, Tri ca17.0101, Tri ca 7, Tri ca 7.0101), Trichostrongylus spp (Tri co 3, Trico 3.0101), Trichophyton spp (Tri eq 4), Trigonella spp (Tri fg 1, Trifg 2, Tri fg 3, Tri fg 4), Trichosanthes spp (Tri k RIP), Trichiurus spp(Tri le 1), Triticum spp (Tri m Peroxidase), Trichophyton spp (Tri me 2,Tri me 4), Trisetum spp (Tri p 1, Tri p 5), Trichinella spp (Tri ps 3,Tri ps 3.0101), Trichophyton spp (Tri r 2, Tri r 2.0101, Tri r 4, Tri r4.0101), Trichoderma spp (Tri rs Cellulase), Triticum spp (Tri s 14),Trichophyton spp (Tri sc 2, Tri sc 4, Tri so 2), Trichinella spp (Tri sp3, Tri sp 3.0101, Tri sp 3.0102, Tri sp 3.0103, Tri sp 3.0104, Tri sp3.0105, Tri sp 3.0106), Trichophyton spp (Tri t 1, Tri t 1.0101, Tri t4, Tri t 4.0101), Triticum spp (Tri td 14, Tri td aA_TI), Trichodermaspp (Tri v Cellulase), Trichophyton spp (Tri ve 4), Triatoma spp (Tria p1, Tria p 1.0101), Triplochiton spp (Trip s 1), Turbo spp (Tur c 1, Turc PM), Tyrophagus spp (Tyr p 1, Tyr p 10, Tyr p 10.0101, Tyr p 10.0102,Tyr p 13, Tyr p 13.0101, Tyr p2, Tyr p 2.0101, Tyr p 24, Tyr p 24.0101,Tyr p 3, Tyr p 3.0101, Tyr p 4, Tyr p 5, Tyr p 5.01, Tyr p 5.02, Tyr p5.03, Tyr p 7, Tyr p alpha Tubulin), Ulocladium spp (Ulo a 1, Ulo at 1,Ulo b 1, Ulo c 1, Ulo co 1, Ulo cu 1, Ulo mu 1, Ulo ob 1, Ulo se 1, Ulosu 1, Ulo to 1), Uncia spp (Unc u 1), Urophycis spp (Uro to 1),Vaccinium spp (Vac m 3), Varroa spp (Var j 13kD), Venerupis spp (Ven ph1, Ven ph 1.0101), Vespula spp (Ves f 1, Ves f 2, Ves f 5, Ves f 5.0101,Ves g 1, Ves g 2, Ves g 5, Ves g 5.0101, Ves m 1, Ves m 1.0101, Ves m 2,Ves m 2.0101, Ves m 5, Ves m 5.0101, Ves m MLT, Ves p 1, Ves p 2, Ves p5, Ves p 5.0101, Ves s 1, Ves s 1.0101, Ves s 2, Ves s 5, Ves s 5.0101,Ves v 1, Ves v 1.0101, Ves v 2, Ves v 2.0101, Ves v 2.0201, Ves v 3, Vesv 3.0101, Ves v 5, Ves v 5.0101, Ves v 5-Pol a 5, Ves vi 5, Ves vi5.0101), Vespa spp (Vesp c 1, Vesp c 1.0101, Vesp c 2, Vesp c 5, Vesp c5.0101, Vesp c 5.0102, Vesp m 1, Vesp m 1.0101, Vesp m 5, Vesp m 5.0101,Vesp ma 1, Vesp ma 2, Vesp ma 5, Vesp ma MLT, Vesp v MLT), Vigna spp(Vig r 1, Vig r 1.0101, Vig r 17kD, Vig r 5, Vig r 8S Globulin, Vig rAlbumin, Vig r beta-Conglycinin), Vitis spp (Vit v 1, Vit v 1.0101, Vitv 4, Vit v 5, Vit v Glucanase, Vit v TLP), Xiphias spp (Xip g 1, Xip g1.0101, Xip g 25kD), Zea spp (Zea m 1, Zea m 1.0101, Zea m 11, Zea m 12,Zea m 12.0101, Zea m 12.0102, Zea m 12.0103, Zea m 12.0104, Zea m12.0105, Zea m 13, Zea m 14, Zea m 14.0101, Zea m 14.0102, Zea m 2, Zeam 20S, Zea m 22, Zea m 25, Zea m 25.0101, Zea m 27kD Zein, Zea m 3, Zeam 4, Zea m 5, Zea m 50kD Zein, Zea m 7, Zea m Chitinase, Zea m G1, Zea mG2, Zea m PAO, Zea m Zml 3), Zeus spp (Zeu fa 1), Ziziphus spp (Ziz m 1,Ziz m 1.0101), Zoarces spp (Zoa a ISP III), Zygophyllum spp (Zyg f 2)

In this context the terms in brackets indicate the particular preferredallergenic antigens (allergens) from the particular source.

Most preferably the allergenic antigen is preferably derived from asource (e.g. a plant (e.g. grass or a tree), a natural product (e.g.milk, nuts etc.), a fungal source (e.g. Aspergillus) or a bacterialsource or from an animal source or animal poison (e.g. cat, dog, venomof bees etc.), preferably selected from the list consisting of grasspollen (e.g. pollen of rye), tree pollen (e.g. pollen of hazel, birch,alder, ash), flower pollen, herb pollen (e.g. pollen of mugwort), dustmite (e.g. Der f 1, Der p 1, Eur m 1, Der m 1 Der f 2, Der p 2, Eur m 2,Tyr p 2, Lep d 2), mold (e.g. allergens of Acremonium, Aspergillus,Cladosporium, Fusarium, Mucor, Penicillium, Rhizopus, Stachybotrys,Trichoderma, or Alternaria), animals (e.g Fel d1, Fel d 2, Fel d3, orFel d4 of cats), food (e.g. allergens of fish (e.g. bass, cod,flounder), seafood (e.g. crab, lobster, shrimps), egg, wheat, nuts (e.g.peanuts, almonds, cashews, walnuts), soya, milk, etc.) or insect venom(e.g. allergens from the venom of wasps, bees, hornets, ants, mosquitos,or ticks).

Autoimmune self-antigens, i.e. antigens associated with autoimmunedisease or autoantigens, may be associated with an autoimmune diseaseaffecting at least one or more of the following organ systems: thecirculatory system, the digestive system, the endocrine system, theexcretory system, the immune system, the integumentary system, themuscular system, the nervous system, the reproductive system, therespiratory system, the skeletal system, preferably with the thecardiovascular system, the neuroendocrine system, the musculoskeletalsystem or gastrointestinal system. Therein the circulatory system is theorgan system which enables pumping and channeling blood to and from thebody and lungs with heart, blood and blood vessels. The digestive systemenables digestion and processing food with salivary glands, esophagus,stomach, liver, gallbladder, pancreas, intestines, colon, rectum andanus. The endocrine system enables communication within the body usinghormones made by endocrine glands such as the hypothalamus, pituitary orpituitary gland, pineal body or pineal gland, thyroid gland, parathyroidgland and adrenal glands. The excretory system comprises kidneys,ureters, bladder and urethra and is involved in fluid balance,electrolyte balance and excretion of urine. The immune system comprisesstructures involved in the transfer of lymph between tissues and theblood stream, the lymph and the nodes and vessels which may beresponsible for transport of cellular and humoral components of theimmune system. It is responsible for defending against disease-causingagents and comprises amongst others leukocytes, tonsils, adenoids,thymus and spleen. The integumentary system comprises skin, hair andnails. The muscular system enables movement with muscles together withthe skeletal system which comprises bones, cartilage, ligaments andtendons and provides structural support. The nervous system isresponsible for collecting, transferring and processing information andcomprises the brain, spinal cord and nerves. The reproductive systemcomprises the sex organs, such as ovaries, fallopian tubes, uterus,vagina, mammary glands, testes, vas deferens, seminal vesicles, prostateand penis. The respiratory system comprises the organs used forbreathing, the pharynx, larynx, trachea, bronchi, lungs and diaphragmand acts together with the circulation system.

Autoimmune self-antigens (antigens associated with autoimmune disease orautoantigens) are selected from autoantigens associated with autoimmunediseases selected from Addison disease (autoimmune adrenalitis, MorbusAddison), alopecia areata, Addison's anemia (Morbus Biermer), autoimmunehemolytic anemia (AIHA), autoimmune hemolytic anemia (AIHA) of the coldtype (cold hemagglutinine disease, cold autoimmune hemolytic anemia(AIHA) (cold agglutinin disease), (CHAD)), autoimmune hemolytic anemia(AIHA) of the warm type (warm AIHA, warm autoimmune haemolytic anemia(AIHA)), autoimmune hemolytic Donath-Landsteiner anemia (paroxysmal coldhemoglobinuria), antiphospholipid syndrome (APS), atherosclerosis,autoimmune arthritis, arteriitis temporalis, Takayasu arteriitis(Takayasu's disease, aortic arch disease), temporal arteriitis/giantcell arteriitis, autoimmune chronic gastritis, autoimmune infertility,autoimmune inner ear disease (AIED), Basedow's disease (Morbus Basedow),Bechterew's disease (Morbus Bechterew, ankylosing spondylitis,spondylitis ankylosans), Behcet's syndrome (Morbus Behcet), boweldisease including autoimmune inflammatory bowel disease (includingcolitis ulcerosa (Morbus Crohn, Crohn's disease), cardiomyopathy,particularly autoimmune cardiomyopathy, idiopathic dilatedcardiomyopathy (DCM), celiac sprue dermatitis (gluten mediatedenteropathia), chronic fatigue immune dysfunction syndrome (CFIDS),chronic inflammatory demyelinating polyneuropathy (CIDP), chronicpolyarthritis, Churg-Strauss syndrome, cicatricial pemphigoid, Cogansyndrome, CREST syndrome (syndrom with Calcinosis cutis, Raynaudphenomenon, motility disorders of the esophagus, sklerodaktylia andteleangiectasia), Crohn's disease (Morbus Crohn, colitis ulcerosa),dermatitis herpetiformis during, dermatologic autoimmune diseases,dermatomyositis, Diabetes, Diabetes mellitus Type 1 (type I diabetes,insuline dependent Diabetes mellitus), Diabetes mellitus Type 2 (type IIdiabetes), essential mixed cryoglobulinemia, essential mixedcryoglobulinemia, fibromyalgia, fibromyositis, Goodpasture syndrome(anti-GBM mediated glomerulonephritis), graft versus host disease,Guillain-Barré syndrome (GBM, Polyradikuloneuritis), haematologicautoimmune diseases, Hashimoto thyroiditis, hemophilia, acquiredhemophilia, hepatitis, autoimmune hepatitis, particularly autoimmuneforms of chronic hepatitis, idiopathic pulmonary fibrosis (IPF),idiopathic thrombocytopenic purpura, Immuno-thrombocytopenic purpura(Morbus Werlhof; ITP), IgA nephropathy, infertility, autoimmuneinfertility, juvenile rheumatoid arthritis (Morbus Still, Stillsyndrome), Lambert-Eaton syndrome, lichen planus, lichen sclerosus,lupus erythematosus, systemic lupus erythematosus (SLE), lupuserythematosus (discoid form), Lyme arthritis (Lyme disease, borreliaarthritis), Ménierè's disease (Morbus Ménierè); mixed connective tissuedisease (MCTD), multiple sclerosis (MS, encephalomyelitis disseminate,Charcot's disease), Myasthenia gravis (myasthenia, MG), myosits,polymyositis, neural autoimmune diseases, neurodermitis, pemphigusvulgaris, bullous pemphigoid, scar forming pemphigoid; polyarteriitisnodosa (periarteiitis nodosa), polychondritis (panchondritis),polyglandular (autoimmune) syndrome (PGA syndrome, Schmidt's syndrome),Polymyalgia rheumatica, primary agammaglobulinemia, primary biliarycirrhosis PBC, primary autoimmune cholangitis), progressive systemicsclerosis (PSS), Psoriasis, Psoriasis vulgaris, Raynaud's phenomena,Reiter's syndrome (Morbus Reiter, urethral conjunctive synovialsyndrome)), rheumatoid arthritis (RA, chronic polyarthritis, rheumaticdisease of the joints, rheumatic fever), sarcoidosis (Morbus Boeck,Besnier-Boeck-Schaumann disease), stiff-man syndrome, Sclerodermia,Scleroderma, Sjögren's syndrome, sympathetic ophtalmia; Transient glutenintolerance, transplanted organ rejection, uveitis, autoimmune uveiitis,Vasculitis, Vitiligo, (leucoderma, piebold skin), and Wegner's disease(Morbus Wegner, Wegner's granulomatosis).

These and other proteins acting as autoimmune self-antigens areunderstood to be therapeutic, as they are meant to treat the subject, inparticular a mammal, more particularly a human being, by vaccinatingwith a self-antigen which is expressed by the mammal, e.g. the human,itself and which triggers an undesired immune response, which is notraised in a healthy subject. Accordingly, such proteins acting asself-antigens are typically of mammalian, in particular human origin.

Particularly preferred in this context are autoimmune self-antigens(autoantigens) selected from:

-   -   myelin basic protein (MBP), proteolipid protein (PLP), and        myelin oligodendrocyte glycoprotein (MOG), in each case        associated with multiple sclerosis (MS);    -   CD44, preproinsulin, proinsulin, insulin, glutamic acid        decaroxylase (GAD65), tyrosine phosphatase-like insulinoma        antigen 2 (IA2), zinc transporter ((ZnT8), and heat shock        protein 60 (HSP60), in each case associated with diabetes Typ I;    -   interphotoreceptor retinoid-binding protein (IRBP) associated        with autoimmune uveitis;    -   acetylcholine receptor AchR, and insulin-like growth factor-1        receptor (IGF-1R), in each case associated with Myasthenia        gravis;    -   M-protein from beta-hemolytic streptocci (pseudo-autoantigen)        associated with Rheumatic Fever;    -   Macrophage migration inhibitory factor associated with        Arthritis;    -   Ro/La RNP complex, alpha- and beta-fodrin, islet cell        autoantigen, poly(ADP)ribose polymerase (PARP), NuMA, NOR-90,        Ro60 autoantigen, and p27 antigen, in each case associated with        Sjögren's syndrome;    -   Ro60 autoantigen, low-density lipoproteins, Sm antigens of the        U-1 small nuclear ribonucleoprotein complex (B/B′, D1, D2, D3,        E, F, G), and RNP ribonucleoproteins, in each case associated        with lupus erythematosus;    -   oxLDL, beta(2)GPI, HSP60/65, and oxLDL/beta(2)GPI, in each case        associated with Atherosclerosis;    -   cardiac beta(1)-adrenergic receptor associated with idiopathic        dilated cardiomyopathy (DCM);    -   histidyl-tRNA synthetase (HisRS) associated with myositis;    -   topoisomerase I associated with scleroderma disease.

Furthermore, in other embodiments said autoimmune self-antigen isassociated with the respective autoimmune disease, like e.g. IL-17, heatshock proteins, and/or any idiotype pathogenic T cell or chemokinereceptor which is expressed by immune cells involved in the autoimmuneresponse in said autoimmune disease (such as any autoimmune diseasesdescribed herein).

The coding region of the inventive nucleic acid according to the firstaspect of the present invention may occur as a mono-, di-, or evenmulticistronic nucleic acid, i.e. a nucleic acid which carries thecoding sequences of one, two or more proteins or peptides. Such codingsequences in di-, or even multicistronic nucleic acids may be separatedby at least one internal ribosome entry site (IRES) sequence, e.g. asdescribed herein or by signal peptides which induce the cleavage of theresulting polypeptide which comprises several proteins or peptides.

According to the first aspect of the present invention, the inventivenucleic acid sequence comprises a coding region, encoding a peptide orprotein which comprises an allergenic antigen or an autoimmuneself-antigen or a fragment, variant or derivative thereof. Preferably,the encoded allergenic antigen or auto-immune self-antigen is no histoneprotein. In the context of the present invention such a histone proteinis typically a strongly alkaline protein found in eukaryotic cellnuclei, which package and order the DNA into structural units callednucleosomes. Histone proteins are the chief protein components ofchromatin, act as spools around which DNA winds, and play a role in generegulation. Without histones, the unwound DNA in chromosomes would bevery long (a length to width ratio of more than 10 million to one inhuman DNA). For example, each human cell has about 1.8 meters of DNA,but wound on the histones it has about 90 millimeters of chromatin,which, when duplicated and condensed during mitosis, result in about 120micrometers of chromosomes. More preferably, in the context of thepresent invention such a histone protein is typically defined as ahighly conserved protein selected from one of the following five majorclasses of histones: H1/H5, H2A, H2B, H3, and H4″, preferably selectedfrom mammalian histone, more preferably from human histones or histoneproteins. Such histones or histone proteins are typically organised intotwo super-classes defined as core histones, comprising histones H2A,H2B, H3 and H4, and linker histones, comprising histones H1 and H5.

In this context, linker histones, preferably excluded from the scope ofprotection of the pending invention, preferably mammalian linkerhistones, more preferably human linker histones, are typically selectedfrom H1, including H1F, particularly including H1F0, H1FNT, H1 FOO, H1FX, and H1H1, particularly including HIST1H1A, HIST1H1B, HIST1H1C,HIST1H1 D, HIST1H1E, HIST1H1T; and

Furthermore, core histones, preferably excluded from the scope ofprotection of the pending invention, preferably mammalian core histones,more preferably human core histones, are typically selected from H2A,including H2AF, particularly including H2AFB1, H2AFB2, H2AFB3, H2AFJ,H2AFV, H2AFX, H2AFY, H2AFY2, H2AFZ, and H2A1, particularly includingHIST1H2AA, HIST1H2AB, HIST1H2AC, HIST1H2AD, HIST1H2AE, HIST1H2AG,HIST1H2A1, HIST1H2AJ, HIST1H2AK, HIST1H2AL, HIST1H2AM, and H2A2,particularly including HIST2H2AA3, HIST2H2AC; H2B, including H2BF,particularly including H2BFM, H2BFO, H2BFS, H2BFWT H2B1, particularlyincluding HIST1H2BA, HIST1H2BB, HIST1H2BC, HIST1H2BD, HIST1H2BE,H1ST1H2BF, HIST1H2BG, HIST1H2BH, HIST1H2BI, HIST1H2BJ, HIST1H2BK,HIST1H2BL, HIST1H2BM, HIST1H2BN, HIST1H2BO, and H2B2, particularlyincluding HIST2H2BE; H3, including H3A1, particularly includingHIST1H3A, HIST1H3B, HIST1H3C, HIST1H3D, HIST1H3E, HIST1H3F, HIST1H3G,HIST1H3H, HIST1H3I, HIST1H3J, and H3A2, particularly including HIST2H3C,and H3A3, particularly including HIST3H3; H4, including H41,particularly including HIST1H4A, HIST1H4B, HIST1H4C, HIST1H4D, HIST1H4E,HIST1H4F, HIST1H4G, HIST1H4H, HIST1H4J, HIST1H4J, HIST1H4K, HIST1H4L,and H44, particularly including HIST4H4, and H5.

According to the first aspect of the present invention, the inventivenucleic acid sequence comprises a coding region, encoding a peptide orprotein which comprises an allergenic antigen or an autoimmuneself-antigen or a fragment, variant or derivative thereof. Preferably,the encoded allergenic antigen or auto-immune self-antigen is noreporter protein (e.g. Luciferase, Green Fluorescent Protein (GFP),Enhanced Green Fluorescent Protein (EGFP), β-Galactosidase) and nomarker or selection protein (e.g. alpha-Globin, Galactokinase andXanthine:guanine phosphoribosyl transferase (GPT)). Preferably, thenucleic acid sequence of the invention does not contain a (bacterial)antibiotics resistance gene, in particular not a neo gene sequence(Neomycin resistance gene) or CAT gene sequence (chloramphenicol acetyltransferase, chloramphenicol resistance gene).

The inventive nucleic acid as define above, comprises or codes for a) acoding region, encoding a peptide or protein which comprises anallergenic antigen or an autoimmune self-antigen or a fragment, variantor derivative thereof; b) at least one histone stem-loop, and c) apoly(A) sequence or polyadenylation signal; preferably for increasingthe expression of said encoded peptide or protein, wherein the encodedpeptide or protein is preferably no histone protein, no reporter proteinand/or no marker or selection protein, as defined above. The elements b)to c) of the inventive nucleic acid may occur in the inventive nucleicacid in any order, i.e. the elements a), b) and c) may occur in theorder a), b) and c) or a), c) and b) from 5′ to 3′ direction in theinventive nucleic acid sequence, wherein further elements as describedherein, may also be contained, such as a 5′-CAP structure, a poly(C)sequence, stabilization sequences, IRES sequences, etc. Each of theelements a) to c) of the inventive nucleic acid, particularly a) in di-or multicistronic constructs and/or each of the elements b) and c), morepreferably element b) may also be repeated at least once, preferablytwice or more in the inventive nucleic acid. As an example, theinventive nucleic acid may show its sequence elements a), b) andoptionally c) in e.g. the following order:

5′-coding region-histone stem-loop-poly(A) sequence-3′; or5′-coding region-histone stem-loop-polyadenylation signal-3′; or5′-coding region-poly(A) sequence-histone stem-loop-3′; or5′-coding region-polyadenylation signal-histone stem-loop-3′; or5′-coding region-coding region-histone stem-loop-polyadenylationsignal-3′; or5′-coding region-histone stem-loop-histone stem-loop-poly(A)sequence-3′; or5′-coding region-histone stem-loop-histone stem-loop-polyadenylationsignal-3′; etc.

In this context it is particularly preferred that the inventive nucleicacid sequence comprises or codes for a) a coding region, encoding apeptide or protein which comprises an allergenic antigen or anautoimmune self-antigen or fragment, variant or derivative thereof; b)at least one histone stem-loop, and c) a poly(A) sequence orpolyadenylation sequence; preferably for increasing the expression levelof said encoded peptide or protein, wherein the encoded protein ispreferably no histone protein, no reporter protein (e.g. Luciferase,GFP, EGFP, β-Galactosidase, particularly EGFP) and/or no marker orselection protein (e.g. alpha-Globin, Galactokinase and Xanthine:Guaninephosphoribosyl transferase (GPT)).

In a further preferred embodiment of the first aspect the inventivenucleic acid sequence as defined herein may also occur in the form of amodified nucleic acid.

In this context, the inventive nucleic acid sequence as defined hereinmay be modified to provide a “stabilized nucleic acid”, preferably astabilized RNA, more preferably an RNA that is essentially resistant toin vivo degradation (e.g. by an exo- or endo-nuclease). A stabilizednucleic acid may e.g. be obtained by modification of the G/C content ofthe coding region of the inventive nucleic acid sequence, byintroduction of nucleotide analogues (e.g. nucleotides with backbonemodifications, sugar modifications or base modifications) or byintroduction of stabilization sequences in the 3′- and/or5′-untranslated region of the inventive nucleic acid sequence.

As mentioned above, the inventive nucleic acid sequence as definedherein may contain nucleotide analogues/modifications e.g. backbonemodifications, sugar modifications or base modifications. A backbonemodification in connection with the present invention is a modificationin which phosphates of the backbone of the nucleotides contained ininventive nucleic acid sequence as defined herein are chemicallymodified. A sugar modification in connection with the present inventionis a chemical modification of the sugar of the nucleotides of theinventive nucleic acid sequence as defined herein. Furthermore, a basemodification in connection with the present invention is a chemicalmodification of the base moiety of the nucleotides of the nucleic acidmolecule of the inventive nucleic acid sequence. In this contextnucleotide analogues or modifications are preferably selected fromnucleotide analogues which are applicable for transcription and/ortranslation.

In a particular preferred embodiment of the first aspect of the presentinvention the herein defined nucleotide analogues/modifications areselected from base modifications which additionally increase theexpression of the encoded protein and which are preferably selected from2-amino-6-chloropurineriboside-5′-triphosphate,2-aminoadenosine-5′-triphosphate, 2-thiocytidine-5′-triphosphate,2-thiouridine-5′-triphosphate, 4-thiouridine-5′-triphosphate,5-aminoallylcytidine-5′-triphosphate,5-aminoallyluridine-5′-triphosphate, 5-bromocytidine-5′-triphosphate,5-bromouridine-5′-triphosphate, 5-iodocytidine-5′-triphosphate,5-iodouridine-5′-triphosphate, 5-methylcytidine-5′-triphosphate,5-methyluridine-5′-triphosphate, 6-azacytidine-5′-triphosphate,6-azauridine-5′-triphosphate, 6-chloropurineriboside-5′-triphosphate,7-deazaadenosine-5′-triphosphate, 7-deazaguanosine-5′-triphosphate,8-azaadenosine-5′-triphosphate, 8-azidoadenosine-5′-triphosphate,benzimidazole-riboside-5′-triphosphate,N1-methyladenosine-5′-triphosphate, N1-methylguanosine-5′-triphosphate,N6-methyladenosine-5′-triphosphate, 06-methylguanosine-5′-triphosphate,pseudouridine-5′-triphosphate, or puromycin-5′-triphosphate,xanthosine-5′-triphosphate. Particular preference is given tonucleotides for base modifications selected from the group ofbase-modified nucleotides consisting of5-methylcytidine-5′-triphosphate, 7-deazaguanosine-5′-triphosphate,5-bromocytidine-5′-triphosphate, and pseudouridine-5′-triphosphate.

According to a further embodiment, the inventive nucleic acid sequenceas defined herein can contain a lipid modification. Such alipid-modified nucleic acid typically comprises a nucleic acid asdefined herein. Such a lipid-modified nucleic acid molecule of theinventive nucleic acid sequence as defined herein typically furthercomprises at least one linker covalently linked with that nucleic acidmolecule, and at least one lipid covalently linked with the respectivelinker. Alternatively, the lipid-modified nucleic acid moleculecomprises at least one nucleic acid molecule as defined herein and atleast one (bifunctional) lipid covalently linked (without a linker) withthat nucleic acid molecule. According to a third alternative, thelipid-modified nucleic acid molecule comprises a nucleic acid moleculeas defined herein, at least one linker covalently linked with thatnucleic acid molecule, and at least one lipid covalently linked with therespective linker, and also at least one (bifunctional) lipid covalentlylinked (without a linker) with that nucleic acid molecule. In thiscontext it is particularly preferred that the lipid modification ispresent at the terminal ends of a linear inventive nucleic acidsequence.

According to another preferred embodiment of the first aspect of theinvention, the inventive nucleic acid sequence as defined herein,particularly if provided as an (m)RNA, can therefore be stabilizedagainst degradation by RNases by the addition of a so-called “5′ CAP”structure.

According to a further preferred embodiment of the first aspect of theinvention, the inventive nucleic acid sequence as defined herein can bemodified by a sequence of at least 10 cytidines, preferably at least 20cytidines, more preferably at least 30 cytidines (so-called “poly(C)sequence”). Particularly, the inventive nucleic acid sequence maycontain or code for a poly(C) sequence of typically about 10 to 200cytidine nucleotides, preferably about 10 to 100 cytidine nucleotides,more preferably about 10 to 70 cytidine nucleotides or even morepreferably about 20 to 50 or even 20 to 30 cytidine nucleotides. Thispoly(C) sequence is preferably located 3′ of the coding region comprisedin the inventive nucleic acid according to the first aspect of thepresent invention.

In a particularly preferred embodiment of the present invention, the G/Ccontent of the coding region, encoding at least one peptide or proteinwhich comprises an allergenic antigen or an autoimmune self-antigen or afragment, variant or derivative thereof of the inventive nucleic acidsequence as defined herein, is modified, particularly increased,compared to the G/C content of its particular wild type coding region,i.e. the unmodified coding region. The encoded amino acid sequence ofthe coding region is preferably not modified compared to the coded aminoacid sequence of the particular wild type coding region.

The modification of the G/C-content of the coding region of theinventive nucleic acid sequence as defined herein is based on the factthat the sequence of any mRNA region to be translated is important forefficient translation of that mRNA. Thus, the composition and thesequence of various nucleotides are important. In particular, mRNAsequences having an increased G (guanosine)/C (cytosine) content aremore stable than mRNA sequences having an increased A (adenosine)/U(uracil) content. According to the invention, the codons of the codingregion are therefore varied compared to its wild type coding region,while retaining the translated amino acid sequence, such that theyinclude an increased amount of G/C nucleotides. In respect to the factthat several codons code for one and the same amino acid (so-calleddegeneration of the genetic code), the most favourable codons for thestability can be determined (so-called alternative codon usage).

Depending on the amino acid to be encoded by the coding region of theinventive nucleic acid sequence as defined herein, there are variouspossibilities for modification of the nucleic acid sequence, e.g. thecoding region, compared to its wild type coding region. In the case ofamino acids which are encoded by codons which contain exclusively G or Cnucleotides, no modification of the codon is necessary. Thus, the codonsfor Pro (CCC or CCG), Arg (CGC or CGG), Ala (GCC or GCG) and Gly (GGC orGGG) require no modification, since no A or U is present.

In contrast, codons which contain A and/or U nucleotides can be modifiedby substitution of other codons which code for the same amino acids butcontain no A and/or U. Examples of these are:

the codons for Pro can be modified from CCU or CCA to CCC or CCG;the codons for Arg can be modified from CGU or CGA or AGA or AGG to CGCor CGG;the codons for Ala can be modified from GCU or GCA to GCC or GCG;the codons for Gly can be modified from GGU or GGA to GGC or GGG.

In other cases, although A or U nucleotides cannot be eliminated fromthe codons, it is however possible to decrease the A and U content byusing codons which contain a lower content of A and/or U nucleotides.Examples of these are:

the codons for Phe can be modified from UUU to UUC;the codons for Leu can be modified from UUA, UUG, CUU or CUA to CUC orCUG;the codons for Ser can be modified from UCU or UCA or AGU to UCC, UCG orAGC;the codon for Tyr can be modified from UAU to UAC;the codon for Cys can be modified from UGU to UGC;the codon for His can be modified from CAU to CAC;the codon for Gln can be modified from CAA to CAG;the codons for Ile can be modified from AUU or AUA to AUC;the codons for Thr can be modified from ACU or ACA to ACC or ACG;the codon for Asn can be modified from AAU to AAC;the codon for Lys can be modified from AAA to AAG;the codons for Val can be modified from GUU or GUA to GUC or GUG;the codon for Asp can be modified from GAU to GAC;the codon for Glu can be modified from GAA to GAG;the stop codon UAA can be modified to UAG or UGA.

In the case of the codons for Met (AUG) and Trp (UGG), on the otherhand, there is no possibility of sequence modification.

The substitutions listed above can be used either individually or in allpossible combinations to increase the G/C content of the coding regionof the inventive nucleic acid sequence as defined herein, compared toits particular wild type coding region (i.e. the original sequence).Thus, for example, all codons for Thr occurring in the wild typesequence can be modified to ACC (or ACG).

In the above context, codons present in mRNA are shown. Thereforeuridine present in an mRNA may also be present as thymidine in therespective DNA coding for the particular mRNA.

Preferably, the G/C content of the coding region of the inventivenucleic acid sequence as defined herein is increased by at least 7%,more preferably by at least 15%, particularly preferably by at least20%, compared to the G/C content of the wild type coding region.According to a specific embodiment at least 5%, 10%, 20%, 30%, 40%, 50%,60%, more preferably at least 70%, even more preferably at least 80% andmost preferably at least 90%, 95% or even 100% of the substitutablecodons in the coding region encoding at least one peptide or proteinwhich comprises an allergenic antigen or an autoimmune self-antigen or afragment, variant or derivative thereof are substituted, therebyincreasing the G/C content of said coding region.

In this context, it is particularly preferable to increase the G/Ccontent of the coding region of the inventive nucleic acid sequence asdefined herein, to the maximum (i.e. 100% of the substitutable codons),compared to the wild type coding region.

According to the invention, a further preferred modification of thecoding region encoding at least one peptide or protein which comprisesan allergenic antigen or an autoimmune self-antigen or a fragment,variant or derivative thereof of the inventive nucleic acid sequence asdefined herein, is based on the finding that the translation efficiencyis also determined by a different frequency in the occurrence of tRNAsin cells. Thus, if so-called “rare codons” are present in the codingregion of the inventive nucleic acid sequence as defined herein, to anincreased extent, the corresponding modified nucleic acid sequence istranslated to a significantly poorer degree than in the case wherecodons coding for relatively “frequent” tRNAs are present.

In this context the coding region of the inventive nucleic acid sequenceis preferably modified compared to the corresponding wild type codingregion such that at least one codon of the wild type sequence whichcodes for a tRNA which is relatively rare in the cell is exchanged for acodon which codes for a tRNA which is relatively frequent in the celland carries the same amino acid as the relatively rare tRNA. By thismodification, the coding region of the inventive nucleic acid sequenceas defined herein, is modified such that codons for which frequentlyoccurring tRNAs are available are inserted. In other words, according tothe invention, by this modification all codons of the wild type codingregion which code for a tRNA which is relatively rare in the cell can ineach case be exchanged for a codon which codes for a tRNA which isrelatively frequent in the cell and which, in each case, carries thesame amino acid as the relatively rare tRNA.

Which tRNAs occur relatively frequently in the cell and which, incontrast, occur relatively rarely is known to a person skilled in theart; cf. e.g. Akashi, Curr. Opin. Genet. Dev. 2001, 11(6): 660-666. Thecodons which use for the particular amino acid the tRNA which occurs themost frequently, e.g. the Gly codon, which uses the tRNA which occursthe most frequently in the (human) cell, are particularly preferred.

According to the invention, it is particularly preferable to link thesequential G/C content which is increased, in particular maximized, inthe coding region of the inventive nucleic acid sequence as definedherein, with the “frequent” codons without modifying the amino acidsequence of the peptide or protein encoded by the coding region of thenucleic acid sequence. This preferred embodiment allows provision of aparticularly efficiently translated and stabilized (modified) inventivenucleic acid sequence as defined herein.

According to another preferred embodiment of the first aspect of theinvention, the inventive nucleic acid sequence as defined herein,preferably has additionally at least one 5′ and/or 3′ stabilizingsequence. These stabilizing sequences in the 5′ and/or 3′ untranslatedregions have the effect of increasing the half-life of the nucleic acid,particularly of the mRNA in the cytosol. These stabilizing sequences canhave 100% sequence identity to naturally occurring sequences which occurin viruses, bacteria and eukaryotes, but can also be partly orcompletely synthetic. The untranslated sequences (UTR) of the(alpha-)globin gene, e.g. from Homo sapiens or Xenopus laevis may bementioned as an example of stabilizing sequences which can be used inthe present invention for a stabilized nucleic acid. Another example ofa stabilizing sequence has the general formula(C/U)CCAN_(x)CCC(U/A)Py_(x)UC(C/U)CC (SEQ ID NO: 55), which is containedin the 3′-UTRs of the very stable RNAs which code for (alpha-)globin,type(I)-collagen, 15-lipoxygenase or for tyrosine hydroxylase (cf.Holcik et al, Proc. Natl. Acad. Sci. USA 1997, 94: 2410 to 2414). Suchstabilizing sequences can of course be used individually or incombination with one another and also in combination with otherstabilizing sequences known to a person skilled in the art. In thiscontext it is particularly preferred that the 3′ UTR sequence of thealpha globin gene is located 3′ of the coding region encoding at leastone peptide or protein which comprises an allergenic antigen or anautoimmune self-antigen or a fragment, variant or derivative thereofcomprised in the inventive nucleic acid sequence according to the firstaspect of the present invention.

Substitutions, additions or eliminations of bases are preferably carriedout with the inventive nucleic acid sequence as defined herein, using aDNA matrix for preparation of the nucleic acid sequence by techniques ofthe well-known site directed mutagenesis or with an oligonucleotideligation strategy (see e.g. Maniatis et al., Molecular Cloning: ALaboratory Manual, Cold Spring Harbor Laboratory Press, 3rd ed., ColdSpring Harbor, N.Y., 2001).

Any of the above modifications may be applied to the inventive nucleicacid sequence as defined herein and further to any nucleic acid as usedin the context of the present invention and may be, if suitable ornecessary, be combined with each other in any combination, provided,these combinations of modifications do not interfere with each other inthe respective nucleic acid. A person skilled in the art will be able totake his choice accordingly.

Nucleic acid sequences used according to the present invention asdefined herein may be prepared using any method known in the art,including synthetic methods such as e.g. solid phase synthesis, as wellas in vitro methods, such as in vitro transcription reactions or in vivoreactions, such as in vivo propagation of DNA plasmids in bacteria.

In such a process, for preparation of the inventive nucleic acidsequence as defined herein, especially if the nucleic acid is in theform of an mRNA, a corresponding DNA molecule may be transcribed invitro. This DNA matrix preferably comprises a suitable promoter, e.g. aT7 or SP6 promoter, for in vitro transcription, which is followed by thedesired nucleotide sequence for the nucleic acid molecule, e.g. mRNA, tobe prepared and a termination signal for in vitro transcription. The DNAmolecule, which forms the matrix of the at least one RNA of interest,may be prepared by fermentative proliferation and subsequent isolationas part of a plasmid which can be replicated in bacteria. Plasmids whichmay be mentioned as suitable for the present invention are e.g. theplasmids pT7 Ts (GenBank accession number U26404; Lai et al.,Development 1995, 121: 2349 to 2360), pGEM® series, e.g. pGEM®-1(GenBank accession number X65300; from Promega) and pSP64 (GenBankaccession number X65327); cf. also Mezei and Storts, Purification of PCRProducts, in: Griffin and Griffin (ed.), PCR Technology: CurrentInnovation, CRC Press, Boca Raton, Fla., 2001.

The inventive nucleic acid sequence as defined herein as well asproteins or peptides as encoded by this nucleic acid sequence maycomprise fragments or variants of those sequences. Such fragments orvariants may typically comprise a sequence having a sequence identitywith one of the above mentioned nucleic acids, or with one of theproteins or peptides or sequences, if encoded by the inventive nucleicacid sequence, of at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, preferablyat least 70%, more preferably at least 80%, equally more preferably atleast 85%, even more preferably at least 90% and most preferably atleast 95% or even 97%, 98% or 99%, to the entire wild type sequence,either on nucleic acid level or on amino acid level.

“Fragments” of proteins or peptides in the context of the presentinvention (e.g. as encoded by the inventive nucleic acid sequence asdefined herein) may comprise a sequence of a protein or peptide asdefined herein, which is, with regard to its amino acid sequence (or itsencoded nucleic acid molecule), N-terminally, C-terminally and/orintrasequentially truncated/shortened compared to the amino acidsequence of the original (native) protein (or its encoded nucleic acidmolecule). Such truncation may thus occur either on the amino acid levelor correspondingly on the nucleic acid level. A sequence identity withrespect to such a fragment as defined herein may therefore preferablyrefer to the entire protein or peptide as defined herein or to theentire (coding) nucleic acid molecule of such a protein or peptide.Likewise, “fragments” of nucleic acids in the context of the presentinvention may comprise a sequence of a nucleic acid as defined herein,which is, with regard to its nucleic acid molecule 5′-, 3′- and/orintrasequentially truncated7shortened compared to the nucleic acidmolecule of the original (native) nucleic acid molecule. A sequenceidentity with respect to such a fragment as defined herein may thereforepreferably refer to the entire nucleic acid as defined herein and thepreferred sequence identity level typically is as indicated herein.Fragments have the same biological function or specific activity or atleast retain an activity of the natural full length protein of at least50%, more preferably at least 70%, even more preferably at least 90%(measured in an appropriate functional assay, e.g. an assay assessingthe immune reaction, e.g. the TH1 and/TH2 reaction by e.g. detecting thesecretion and/or expression of cytokines indicating the strength of theimmune response) as compared to the full-length native wt peptide orprotein, e.g. its specific antigenic or therapeutic property.Accordingly, in a preferred embodiment, the “fragment” is a portion ofthe full-length (naturally occurring) wt allergenic or self-antigenicprotein, which exerts therapeutic properties as indicated herein, e.g.for vaccination.

Fragments of proteins or peptides in the context of the presentinvention (e.g. as encoded by the inventive nucleic acid sequence asdefined herein) may furthermore comprise a sequence of a protein orpeptide as defined herein, which has a length of about 6 to about 20 oreven more amino acids, e.g. fragments as processed and presented by MHCclass I molecules, preferably having a length of about 8 to about 10amino acids, e.g. 8, 9, or 10, (or even 6, 7, 11, or 12 amino acids), orfragments as processed and presented by MHC class II molecules,preferably having a length of about 13 or more amino acids, e.g. 13, 14,15, 16, 17, 18, 19, 20 or even more amino acids, wherein these fragmentsmay be selected from any part of the amino acid sequence. Thesefragments are typically recognized by T-cells in form of a complexconsisting of the peptide fragment and an MHC molecule, i.e. thefragments are typically not recognized in their native form. Fragmentsof proteins or peptides as defined herein may comprise at least oneepitope of those proteins or peptides. Furthermore also domains of aprotein, like the extracellular domain, the intracellular domain or thetransmembrane domain and shortened or truncated versions of a proteinmay be understood to comprise/correspond to a fragment of a protein.

Fragments of proteins or peptides as defined herein (e.g. as encoded bythe inventive nucleic acid sequence as defined herein) may also compriseepitopes of those proteins or peptides. T cell epitopes or parts of theproteins in the context of the present invention may comprise fragmentspreferably having a length of about 6 to about 20 or even more aminoacids, e.g. fragments as processed and presented by MHC class Imolecules, preferably having a length of about 8 to about 10 aminoacids, e.g. 8, 9, or 10, (or even 11, or 12 amino acids), or fragmentsas processed and presented by MHC class II molecules, preferably havinga length of about 13 or more amino acids, e.g. 13, 14, 15, 16, 17, 18,19, 20 or even more amino acids, wherein these fragments may be selectedfrom any part of the amino acid sequence. These fragments are typicallyrecognized by T cells in form of a complex consisting of the peptidefragment and an MHC molecule, i.e. the fragments are typically notrecognized in their native form.

B cell epitopes are typically fragments located on the outer surface of(native) protein or peptide antigens as defined herein, preferablyhaving 5 to 15 amino acids, more preferably having 5 to 12 amino acids,even more preferably having 6 to 9 amino acids, which may be recognizedby antibodies, i.e. in their native form.

Such epitopes of proteins or peptides may furthermore be selected fromany of the herein mentioned variants of such proteins or peptides. Inthis context antigenic determinants can be conformational ordiscontinuous epitopes which are composed of segments of the proteins orpeptides as defined herein that are discontinuous in the amino acidsequence of the proteins or peptides as defined herein but are broughttogether in the three-dimensional structure or continuous or linearepitopes which are composed of a single polypeptide chain.

“Variants” of proteins or peptides as defined in the context of thepresent invention may be encoded by the inventive nucleic acid sequenceas defined herein. Thereby, a protein or peptide may be generated,having an amino acid sequence which differs from the original sequencein one or more (2, 3, 4, 5, 6, 7 or more) mutation(s), such as one ormore substituted, inserted and/or deleted amino acid(s). The preferredlevel of sequence identity of “variants” in view of the full-lengthnatural wt protein sequence typically is as indicated herein.Preferably, variants have the same biological function or specificactivity or at least retain an activity of the natural full lengthprotein of at least 50%, more preferably at least 70%, even morepreferably at least 90% (measured in an appropriate functional assay,e.g. an assay assessing the immune reaction, e.g. the TH1 and/TH2reaction by e.g. detecting the secretion and/or expression of cytokinesindicating the strength of the immune response) as compared to thefull-length native peptide or protein, e.g. its specific antigenic oftherapeutic property. Accordingly, in a preferred embodiment, the“variant” is a variant of an allergenic or self-antigenic protein, whichexerts therapeutic properties, e.g. as a vaccine, to the extent asindicated herein.

“Variants” of proteins or peptides as defined in the context of thepresent invention (e.g. as encoded by a nucleic acid as defined herein)may comprise conservative amino acid substitution(s) compared to theirnative, i.e. non-mutated physiological, sequence. Those amino acidsequences as well as their encoding nucleotide sequences in particularfall under the term variants as defined herein. Substitutions in whichamino acids, which originate from the same class, are exchanged for oneanother are called conservative substitutions. In particular, these areamino acids having aliphatic side chains, positively or negativelycharged side chains, aromatic groups in the side chains or amino acids,the side chains of which can enter into hydrogen bridges, e.g. sidechains which have a hydroxyl function. This means that e.g. an aminoacid having a polar side chain is replaced by another amino acid havinga likewise polar side chain, or, for example, an amino acidcharacterized by a hydrophobic side chain is substituted by anotheramino acid having a likewise hydrophobic side chain (e.g. serine(threonine) by threonine (serine) or leucine (isoleucine) by isoleucine(leucine)). Insertions and substitutions are possible, in particular, atthose sequence positions which cause no modification to thethree-dimensional structure or do not affect the binding region.Modifications to a three-dimensional structure by insertion(s) ordeletion(s) can easily be determined e.g. using CD spectra (circulardichroism spectra) (Urry, 1985, Absorption, Circular Dichroism and ORDof Polypeptides, in: Modern Physical Methods in Biochemistry, Neubergeret al. (ed.), Elsevier, Amsterdam).

Furthermore, variants of proteins or peptides as defined herein, whichmay be encoded by the inventive nucleic acid sequence as defined herein,may also comprise those sequences, wherein nucleotides of the nucleicacid are exchanged according to the degeneration of the genetic code,without leading to an alteration of the respective amino acid sequenceof the protein or peptide, i.e. the amino acid sequence or at least partthereof may not differ from the original sequence in one or moremutation(s) within the above meaning.

In order to determine the percentage to which two sequences areidentical, e.g. nucleic acid sequences or amino acid sequences asdefined herein, preferably the amino acid sequences encoded by theinventive nucleic acid sequence as defined herein or the amino acidsequences themselves, the sequences can be aligned in order to besubsequently compared to one another. Therefore, e.g. a position of afirst sequence may be compared with the corresponding position of thesecond sequence. If a position in the first sequence is occupied by thesame component as is the case at a position in the second sequence, thetwo sequences are identical at this position. If this is not the case,the sequences differ at this position. If insertions occur in the secondsequence in comparison to the first sequence, gaps can be inserted intothe first sequence to allow a further alignment. If deletions occur inthe second sequence in comparison to the first sequence, gaps can beinserted into the second sequence to allow a further alignment. Thepercentage to which two sequences are identical is then a function ofthe number of identical positions divided by the total number ofpositions including those positions which are only occupied in onesequence. The percentage to which two sequences are identical can bedetermined using a mathematical algorithm. A preferred, but notlimiting, example of a mathematical algorithm which can be used is thealgorithm of Karlin et al. (1993), PNAS USA, 90:5873-5877 or Altschul etal. (1997), Nucleic Acids Res., 25:3389-3402. Such an algorithm isintegrated in the BLAST program. Sequences which are identical to thesequences of the present invention to a certain extent can be identifiedby this program.

The inventive nucleic acid sequence as defined herein may encodederivatives of a peptide or protein. Such a derivative of a peptide orprotein is a molecule that is derived from another molecule, such assaid peptide or protein. A “derivative” typically contains thefull-length sequence of the natural wt peptide or protein and additionalsequence features, e.g. at the N- or at the C-terminus, which mayexhibit an additional function to the natural full-lengthpeptide/protein. Again such derivatives have the same biologicalfunction or specific activity or at least retain an activity of thenatural wt full-length protein of at least 50%, more preferably at least70%, even more preferably at least 90% (measured in an appropriatefunctional assay, see above, e.g. an assay measuring the immunereaction) as compared to the full-length native wt peptide or protein,e.g. its specific allergenic/self-antigenic therapeutic property.Thereby, a “derivative” of a peptide or protein also encompasses(chimeric) fusion peptides/proteins comprising a peptide or protein usedin the present invention or a natural wt full-length protein (orvariant/fragment thereof) fused to a distinct peptide/protein awardinge.g. two or more biological functions to the fusion peptide/protein. Forexample, the fusion may comprise a label, such as, for example, anepitope, e.g., a FLAG epitope or a V5 epitope or an HA epitope. Forexample, the epitope is a FLAG epitope. Such a tag is useful for, forexample, purifying the fusion protein.

In this context, a “variant” of a protein or peptide may have at least70%, 75%, 80%, 85%, 90%, 95%, 98% or 99% amino acid identity over astretch of 10, 20, 30, 50, 75 or 100 amino acids of such protein orpeptide. Analogously, a “variant” of a nucleic acid sequence or,particularly, a fragment, may have at least 70%, 75%, 80%, 85%, 90%,95%, 98% or 99% nucleotide identity over a stretch of 10, 20, 30, 50, 75or 100 nucleotide of such nucleic acid sequence; typically, however,referring to the naturally occurring wt full-length sequences. In caseof “fragments” typically, sequence identity is determined for thefragment over the length (of the fragment) on the portion of thefull-length protein (reflecting the same length as the fragment), whichexhibits the highest level of sequence identity.

In a further preferred embodiment of the first aspect of the presentinvention the inventive nucleic acid sequence is associated with avehicle, transfection or complexation agent for increasing thetransfection efficiency and/or the immunostimulatory properties of theinventive nucleic acid sequence. Particularly preferred agents in thiscontext suitable for increasing the transfection efficiency are cationicor polycationic compounds, including protamine, nucleoline, spermine orspermidine, or other cationic peptides or proteins, such aspoly-L-lysine (PLL), poly-arginine, basic polypeptides, cell penetratingpeptides (CPPs), including HIV-binding peptides, HIV-1 Tat (HIV),Tat-derived peptides, Penetratin, VP22 derived or analog peptides, HSVVP22 (Herpes simplex), MAP, KALA or protein transduction domains (PTDs),PpT620, prolin-rich peptides, arginine-rich peptides, lysine-richpeptides, MPG-peptide(s), Pep-1, L-oligomers, Calcitonin peptide(s),Antennapedia-derived peptides (particularly from Drosophilaantennapedia), pAntp, pIsl, FGF, Lactoferrin, Transportan, Buforin-2,Bac715-24, SynB, SynB(1), pVEC, hCT-derived peptides, SAP, or histones.Additionally, preferred cationic or polycationic proteins or peptidesmay be selected from the following proteins or peptides having thefollowing total formula:(Arg)_(i);(Lys)_(m);(His)_(n);(Orn)_(o);(Xaa)_(x), whereinI+m+n+o+x=8-15, and l, m, n or o independently of each other may be anynumber selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or15, provided that the overall content of Arg, Lys, His and Ornrepresents at least 50% of all amino acids of the oligopeptide; and Xaamay be any amino acid selected from native (=naturally occurring) ornon-native amino acids except of Arg, Lys, His or Orn; and x may be anynumber selected from 0, 1, 2, 3 or 4, provided, that the overall contentof Xaa does not exceed 50% of all amino acids of the oligopeptide.Particularly preferred cationic peptides in this context are e.g. Arg₇,Arg₈, Arg₉, H₃R₉, R₉H₃, H₃R₉H₃, YSSR₉SSY, (RKH)₄, Y(RKH)₂R, etc. Furtherpreferred cationic or polycationic compounds, which can be used astransfection agent may include cationic polysaccharides, for examplechitosan, polybrene, cationic polymers, e.g. polyethyleneimine (PEI),cationic lipids, e.g. DOTMA:[1-(2,3-sioleyloxy)propyl)]-N,N,N-trimethylammonium chloride, DMRIE,di-C14-amidine, DOTIM, SAINT, DC-Chol, BGTC, CTAP, DOPC, DODAP, DOPE:Dioleyl phosphatidylethanol-amine, DOSPA, DODAB, DOIC, DMEPC, DOGS:Dioctadecylamidoglicylspermin, DIMRI: Dimyristo-oxypropyl dimethylhydroxyethyl ammonium bromide, DOTAP:dioleoyloxy-3-(trimethylammonio)propane, DC-6-14:0,0-ditetradecanoyl-N-(a-trimethylammonioacetyl)diethanolamine chloride,CLIP1: rac-[(2,3-dioctadecyloxypropyl)(2-hydroxyethyl)]-dimethylammoniumchloride, CLIP6:rac-[2(2,3-dihexadecyloxypropyl-oxymethyloxy)ethyl]trimethylammonium,CLIP9:rac-[2(2,3-dihexadecyloxypropyl-oxysuccinyloxy)ethyl]-trimethylammonium,oligofectamine, or cationic or polycationic polymers, e.g. modifiedpolyaminoacids, such as β-aminoacid-polymers or reversed polyamides,etc., modified polyethylenes, such as PVP(poly(N-ethyl-4-vinylpyridinium bromide)), etc., modified acrylates,such as pDMAEMA (poly(dimethylaminoethyl methylacrylate)), etc.,modified Amidoamines such as pAMAM (poly(amidoamine)), etc., modifiedpolybetaaminoester (PBAE), such as diamine end modified 1,4 butanedioldiacrylate-co-5-amino-1-pentanol polymers, etc., dendrimers, such aspolypropylamine dendrimers or pAMAM based dendrimers, etc.,polyimine(s), such as PEI: poly(ethyleneimine), poly(propyleneimine),etc., polyallylamine, sugar backbone based polymers, such ascyclodextrin based polymers, dextran based polymers, chitosan, etc.,silan backbone based polymers, such as PMOXA-PDMS copolymers, etc.,blockpolymers consisting of a combination of one or more cationic blocks(e.g. selected from a cationic polymer as mentioned above) and of one ormore hydrophilic or hydrophobic blocks (e.g polyethyleneglycole); etc.

In this context it is particularly preferred that the inventive nucleicacid is complexed at least partially with a cationic or polycationiccompound, preferably cationic proteins or peptides. Partially means thatonly a part of the inventive nucleic acid is complexed with a cationicor polycationic compound and that the rest of the inventive nucleic acidis in uncomplexed form (“free”). Preferably the ratio of complexednucleic acid to:free nucleic acid is selected from a range. of about 5:1(w/w) to about 1:10 (w/w), more preferably from a range of about 4:1(w/w) to about 1:8 (w/w), even more preferably from a range of about 3:1(w/w) to about 1:5 (w/w) or 1:3 (w/w), and most preferably the ratio ofcomplexed nucleic acid to free nucleic acid is selected from a ratio ofabout 1:1 (w/w).

It is preferred that the nucleic acid sequence of the invention isprovided in either naked form or complexed, e.g. by polycationiccompounds of whatever chemical structure, preferably polycationic(poly)peptides or synthetic polycationic compounds. Preferably, thenucleic acid sequence is not provided together with a packaging cell.

In a further aspect the invention provides for a composition or kit orkit of parts comprising a plurality or more than one, preferably 2 to10, more preferably 2 to 5, most preferably 2 to 4 of the inventivenucleic acid sequences as defined herein. These inventive compositionscomprise more than one inventive nucleic acid sequences, preferablyencoding different peptides or proteins which comprise preferablydifferent allergenic antigens or auto-immune self-antigens or fragments,variants or derivatives thereof.

According to a further aspect, the present invention also provides amethod for increasing the expression of an encoded peptide or proteincomprising the steps, e.g. a) providing the inventive nucleic acidsequence as defined herein or the inventive composition comprising aplurality (which means typically more than 1, 2, 3, 4, 5, 6 or more than10 nucleic acids, e.g. 2 to 10, preferably 2 to 5 nucleic acids) ofinventive nucleic acid sequences as defined herein, b) applying oradministering the inventive nucleic acid sequence as defined herein orthe inventive composition comprising a plurality of inventive nucleicacid sequences as defined herein to an expression system, e.g. to acell-free expression system, a cell (e.g. an expression host cell or asomatic cell), a tissue or an organism. The method may be applied forlaboratory, for research, for diagnostic, for commercial production ofpeptides or proteins and/or for therapeutic purposes. In this context,typically after preparing the inventive nucleic acid sequence as definedherein or of the inventive composition comprising a plurality ofinventive nucleic acid sequences as defined herein, it is typicallyapplied or administered to a cell-free expression system, a cell (e.g.an expression host cell or a somatic cell), a tissue or an organism,e.g. in naked or complexed form or as a pharmaceutical composition orvaccine as described herein, preferably via transfection or by using anyof the administration modes as described herein. The method may becarried out in vitro, in vivo or ex vivo. The method may furthermore becarried out in the context of the treatment of a specific disease,particularly in the treatment of allergies or autoimmune diseases,preferably as defined herein.

In this context in vitro is defined herein as transfection ortransduction of the inventive nucleic acid as defined herein or of theinventive composition comprising a plurality of inventive nucleic acidsequences as defined herein into cells in culture outside of anorganism; in vivo is defined herein as transfection or transduction ofthe inventive nucleic acid or of the inventive composition comprising aplurality of inventive nucleic acid sequences (which means typicallymore than 1, 2, 3, 4, 5, 6 or more than 10 nucleic acids, e.g. 2 to 10,preferably 2 to 5 nucleic acids) into cells by application of theinventive nucleic acid or of the inventive composition to the wholeorganism or individual and ex vivo is defined herein as transfection ortransduction of the inventive nucleic acid or of the inventivecomposition comprising a plurality of inventive nucleic acid sequencesinto cells outside of an organism or individual and subsequentapplication of the transfected cells to the organism or individual.

Likewise, according to another aspect, the present invention alsoprovides the use of the inventive nucleic acid sequence as definedherein or of the inventive composition comprising a plurality ofinventive nucleic acid sequences as defined herein, preferably fordiagnostic or therapeutic purposes, for increasing the expression of anencoded peptide or protein, e.g. by applying or administering theinventive nucleic acid sequence as defined herein or of the inventivecomposition comprising a plurality of inventive nucleic acid sequencesas defined herein, e.g. to a cell-free expression system, a cell (e.g.an expression host cell or a somatic cell), a tissue or an organism. Theuse may be applied for laboratory, for research, for diagnostic forcommercial production of peptides or proteins and/or for therapeuticpurposes. In this context, typically after preparing the inventivenucleic acid sequence as defined herein or of the inventive compositioncomprising a plurality of inventive nucleic acid sequences as definedherein, it is typically applied or administered to a cell-freeexpression system, a cell (e.g. an expression host cell or a somaticcell), a tissue or an organism, preferably in naked form or complexedform, or as a pharmaceutical composition or vaccine as described herein,preferably via transfection or by using any of the administration modesas described herein. The use may be carried out in vitro, in vivo or exvivo. The use may furthermore be carried out in the context of thetreatment of a specific disease, particularly in the treatment ofallergies or autoimmune diseases, preferably as defined herein.

In yet another aspect the present invention also relates to an inventiveexpression system comprising an inventive nucleic acid sequence orexpression vector or plasmid according to the first aspect of thepresent invention. In this context the expression system may be acell-free expression system (e.g. an in vitro transcription/translationsystem), a cellular expression system (e.g. mammalian cells like CHOcells, insect cells, yeast cells, bacterial cells like E. coli) ororganisms used for expression of peptides or proteins (e.g. plants oranimals like cows).

Additionally, according to another aspect, the present invention alsorelates to the use of the inventive nucleic acid as defined herein or ofthe inventive composition comprising a plurality of inventive nucleicacid sequences as defined herein for the preparation of a pharmaceuticalcomposition for increasing the expression of an encoded peptide orprotein, e.g. for treating allergies or autoimmune diseases, preferablyas defined herein, e.g. applying or administering the inventive nucleicacid as defined herein or of the inventive composition comprising aplurality of inventive nucleic acid sequences as defined herein to acell (e.g. an expression host cell or a somatic cell), a tissue or anorganism, preferably in naked form or complexed form or as apharmaceutical composition or vaccine as described herein, morepreferably using any of the administration modes as described herein.

Accordingly, in a particular preferred aspect, the present inventionalso provides a pharmaceutical composition, comprising an inventivenucleic acid as defined herein or an inventive composition comprising aplurality of inventive nucleic acid sequences as defined herein andoptionally a pharmaceutically acceptable carrier and/or vehicle.

As a first ingredient, the inventive pharmaceutical compositioncomprises at least one inventive nucleic acid as defined herein.

As a second ingredient the inventive pharmaceutical composition mayoptional comprise at least one additional pharmaceutically activecomponent. A pharmaceutically active component in this connection is acompound that has a therapeutic effect to heal, ameliorate or prevent aparticular indication or disease as mentioned herein, preferablyallergies or autoimmune diseases. Such compounds include, withoutimplying any limitation, peptides or proteins, preferably as definedherein, nucleic acids, preferably as defined herein, (therapeuticallyactive) low molecular weight organic or inorganic compounds (molecularweight less than 5000, preferably less than 1000), sugars, antigens orantibodies, preferably as defined herein, therapeutic agents alreadyknown in the prior art, antigenic cells, antigenic cellular fragments,cellular fractions; cell wall components (e.g. polysaccharides),modified, attenuated or de-activated (e.g. chemically or by irradiation)pathogens (virus, bacteria etc.), adjuvants, preferably as definedherein, etc.

Furthermore, the inventive pharmaceutical composition may comprise apharmaceutically acceptable carrier and/or vehicle. In the context ofthe present invention, a pharmaceutically acceptable carrier typicallyincludes the liquid or non-liquid basis of the inventive pharmaceuticalcomposition. If the inventive pharmaceutical composition is provided inliquid form, the carrier will typically be pyrogen-free water; isotonicsaline or buffered (aqueous) solutions, e.g phosphate, citrate etc.buffered solutions. The injection buffer may be hypertonic, isotonic orhypotonic with reference to the specific reference medium, i.e. thebuffer may have a higher, identical or lower salt content with referenceto the specific reference medium, wherein preferably such concentrationsof the afore mentioned salts may be used, which do not lead to damage ofcells due to osmosis or other concentration effects. Reference media aree.g. liquids occurring in “in vivo” methods, such as blood, lymph,cytosolic liquids, or other body liquids, or e.g. liquids, which may beused as reference media in “in vitro” methods, such as common buffers orliquids. Such common buffers or liquids are known to a skilled person.Ringer-Lactate solution is particularly preferred as a liquid basis.

However, one or more compatible solid or liquid fillers or diluents orencapsulating compounds may be used as well for the inventivepharmaceutical composition, which are suitable for administration to apatient to be treated. The term “compatible” as used here means thatthese constituents of the inventive pharmaceutical composition arecapable of being mixed with the inventive nucleic acid as defined hereinin such a manner that no interaction occurs which would substantiallyreduce the pharmaceutical effectiveness of the inventive pharmaceuticalcomposition under typical use conditions.

According to a specific embodiment, the inventive pharmaceuticalcomposition may comprise an adjuvant. In this context, an adjuvant maybe understood as any compound, which is suitable to initiate or increasean immune response of the innate immune system, i.e. a non-specificimmune response. With other words, when administered, the inventivepharmaceutical composition preferably elicits an innate immune responsedue to the adjuvant, optionally contained therein. Preferably, such anadjuvant may be selected from an adjuvant known to a skilled person andsuitable for the present case, i.e. supporting the induction of aninnate immune response in a mammal, e.g. an adjuvant protein as definedabove or an adjuvant as defined in the following.

Particularly preferred as adjuvants suitable for depot and delivery arecationic or polycationic compounds as defined above for the inventivenucleic acid sequence as vehicle, transfection or complexation agent.

The inventive pharmaceutical composition can additionally contain one ormore auxiliary substances in order to increase its immunogenicity orimmunostimulatory capacity, if desired. A synergistic action of theinventive nucleic acid sequence as defined herein and of an auxiliarysubstance, which may be optionally contained in the inventivepharmaceutical composition, is preferably achieved thereby. Depending onthe various types of auxiliary substances, various mechanisms can comeinto consideration in this respect. For example, compounds that permitthe maturation of dendritic cells (DCs), for examplelipopolysaccharides, TNF-alpha or CD40 ligand, form a first class ofsuitable auxiliary substances. In general, it is possible to use asauxiliary substance any agent that influences the immune system in themanner of a “danger signal” (LPS, GP96, etc.) or cytokines, such asGM-CFS, which allow an immune response to be enhanced and/or influencedin a targeted manner. Particularly preferred auxiliary substances arecytokines, such as monokines, lymphokines, interleukins or chemokines,that further promote the innate immune response, such as IL-1, IL-2,IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-14,IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, IL-21, IL-22, IL-23, IL-24,IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33,IFN-alpha, IFN-beta, IFN-gamma, GM-CSF, G-CSF, M-CSF, LT-beta orTNF-alpha, growth factors, such as hGH.

Further additives which may be included in the inventive pharmaceuticalcomposition are emulsifiers, such as, for example, Tween®; wettingagents, such as, for example, sodium lauryl sulfate; colouring agents;taste-imparting agents, pharmaceutical carriers; tablet-forming agents;stabilizers; antioxidants; preservatives.

The inventive pharmaceutical composition can also additionally containany further compound, which is known to be immunostimulating due to itsbinding affinity (as ligands) to human Toll-like receptors TLR1, TLR2,TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, or due to its bindingaffinity (as ligands) to murine Toll-like receptors TLR1, TLR2, TLR3,TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, TLR11, TLR12 or TLR13.

The inventive pharmaceutical composition may be administered orally,parenterally, by inhalation spray, topically, rectally, nasally,buccally, vaginally or via an implanted reservoir. The term parenteralas used herein includes subcutaneous, intravenous, intramuscular,intra-articular, intra-synovial, intrasternal, intrathecal,intrahepatic, intralesional, intracranial, transdermal, intradermal,intrapulmonal, intraperitoneal, intracardial, intraarterial, andsublingual injection or infusion techniques.

Preferably, the inventive pharmaceutical composition may be administeredby parenteral injection, more preferably by subcutaneous, intravenous,intramuscular, intra-articular, intra-synovial, intrasternal,intrathecal, intrahepatic, intralesional, intracranial, transdermal,intradermal, intrapulmonal, intraperitoneal, intracardial,intraarterial, and sublingual injection or via infusion techniques.Particularly preferred is intradermal and intramuscular injection.Sterile injectable forms of the inventive pharmaceutical compositionsmay be aqueous or oleaginous suspension. These suspensions may beformulated according to techniques known in the art using suitabledispersing or wetting agents and suspending agents.

The inventive pharmaceutical composition as defined herein may also beadministered orally in any orally acceptable dosage form including, butnot limited to, capsules, tablets, aqueous suspensions or solutions.

The inventive pharmaceutical composition may also be administeredtopically, especially when the target of treatment includes areas ororgans readily accessible by topical application, e.g. includingdiseases of the skin or of any other accessible epithelial tissue.Suitable topical formulations are readily prepared for each of theseareas or organs. For topical applications, the inventive pharmaceuticalcomposition may be formulated in a suitable ointment, containing theinventive nucleic acid as defined herein suspended or dissolved in oneor more carriers.

The inventive pharmaceutical composition typically comprises a “safe andeffective amount” of the components of the inventive pharmaceuticalcomposition, particularly of the inventive nucleic acid sequence(s) asdefined herein. As used herein, a “safe and effective amount” means anamount of the inventive nucleic acid sequence(s) as defined herein assuch that is sufficient to significantly induce a positive modificationof a disease or disorder as defined herein. At the same time, however, a“safe and effective amount” is small enough to avoid seriousside-effects and to permit a sensible relationship between advantage andrisk. The determination of these limits typically lies within the scopeof sensible medical judgment.

The inventive pharmaceutical composition may be used for human and alsofor veterinary medical purposes, preferably for human medical purposes,as a pharmaceutical composition in general or as a vaccine.

According to another particularly preferred aspect, the inventivepharmaceutical composition (or the inventive nucleic acid sequence asdefined herein or the inventive composition comprising a plurality ofinventive nucleic acid sequences as defined herein) may be provided orused as a vaccine. Typically, such a vaccine is as defined above forpharmaceutical compositions. Additionally, such a vaccine typicallycontains the inventive nucleic acid as defined herein or the inventivecomposition comprising a plurality of inventive nucleic acid sequencesas defined herein.

The inventive vaccine may also comprise a pharmaceutically acceptablecarrier, adjuvant, and/or vehicle as defined herein for the inventivepharmaceutical composition. In the specific context of the inventivevaccine, the choice of a pharmaceutically acceptable carrier isdetermined in principle by the manner in which the inventive vaccine isadministered. The inventive vaccine can be administered, for example,systemically or locally. Routes for systemic administration in generalinclude, for example, transdermal, oral, parenteral routes, includingsubcutaneous, intravenous, intramuscular, intraarterial, intradermal andintraperitoneal injections and/or intranasal administration routes.Routes for local administration in general include, for example, topicaladministration routes but also intradermal, transdermal, subcutaneous,or intramuscular injections or intralesional, intracranial,intrapulmonal, intracardial, and sublingual injections. More preferably,vaccines may be administered by an intradermal, subcutaneous, orintramuscular route. Inventive vaccines are therefore preferablyformulated in liquid (or sometimes in solid) form.

The inventive vaccine can additionally contain one or more auxiliarysubstances in order to increase its immunogenicity or immunostimulatorycapacity, if desired. Particularly preferred are adjuvants as auxiliarysubstances or additives as defined for the pharmaceutical composition.

The present invention furthermore provides several applications and usesof the inventive nucleic acid sequence as defined herein, of theinventive composition comprising a plurality of inventive nucleic acidsequences as defined herein, of the inventive pharmaceuticalcomposition, of the inventive vaccine, all comprising the inventivenucleic acid sequence as defined herein or of kits comprising same.

According to one specific aspect, the present invention is directed tothe first medical use of the inventive nucleic acid sequence as definedherein or of the inventive composition comprising a plurality ofinventive nucleic acid sequences as defined herein as a medicament,preferably as a vaccine particularly in the treatment of allergies orautoimmune diseases.

According to another aspect, the present invention is directed to thesecond medical use of the inventive nucleic acid sequence as definedherein or of the inventive composition comprising a plurality ofinventive nucleic acid sequences as defined herein, for the treatmentallergies our autoimmune diseases as defined herein, preferably to theuse of the inventive nucleic acid sequence as defined herein, of theinventive composition comprising a plurality of inventive nucleic acidsequences as defined herein, of a pharmaceutical composition or vaccinecomprising same or of kits comprising same for the preparation of amedicament for the prophylaxis, treatment and/or amelioration ofallergies or autoimmune diseases as defined herein. Preferably, thepharmaceutical composition or a vaccine is used or to be administered toa patient in need thereof for this purpose.

Preferably, diseases as mentioned herein are selected from allergieswhich preferably include e.g. pollen allergy (allergy against grasspollen, tree pollen (e.g. pollen of hazel, birch, alder, ash), flowerpollen, herb pollen (e.g. pollen of mugwort)), dust mite allergy, moldallergy (e.g. allergy against Acremonium, Aspergillus, Cladosporium,Fusarium, Mucor, Penicillium, Rhizopus, Stachybotrys, Trichoderma, orAlternaria), pet allergy (allergy against animals; e.g against cats,dogs, horses), food allergy (e.g. allergy against fish (e.g. bass, cod,flounder), seafood (e.g. crab, lobster, shrimps), egg, wheat, nuts (e.g.peanuts, almonds, cashews, walnuts), soya, milk, etc.) or insect biteallergy (allergy against insect venom, e.g. venom of wasps, bees,hornets, ants, mosquitos, or ticks).

According to another specific embodiment, diseases as defined hereincomprise autoimmune diseases as defined in the following. autoimmunediseases are preferably selected from Addison disease (autoimmuneadrenalitis, Morbus Addison), alopecia areata, Addison's anemia (MorbusBiermer), autoimmune hemolytic anemia (AIHA), autoimmune hemolyticanemia (AIHA) of the cold type (cold hemagglutinine disease, coldautoimmune hemolytic anemia (AIHA) (cold agglutinin disease), (CHAD)),autoimmune hemolytic anemia (AIHA) of the warm type (warm AIHA, warmautoimmune haemolytic anemia (AIHA)), autoimmune hemolyticDonath-Landsteiner anemia (paroxysmal cold hemoglobinuria),antiphospholipid syndrome (APS), atherosclerosis, autoimmune arthritis,arteriitis temporalis, Takayasu arteriitis (Takayasu's disease, aorticarch disease), temporal arteriitis/giant cell arteriitis, autoimmunechronic gastritis, autoimmune infertility, autoimmune inner ear disease(AIED), Basedow's disease (Morbus Basedow), Bechterew's disease (MorbusBechterew, ankylosing spondylitis, spondylitis ankylosans), Behcet'ssyndrome (Morbus Behcet), bowel disease including autoimmuneinflammatory bowel disease (including colitis ulcerosa (Morbus Crohn,Crohn's disease), cardiomyopathy, particularly autoimmunecardiomyopathy, idiopathic dilated cardiomyopathy (DCM), celiac spruedermatitis (gluten mediated enteropathia), chronic fatigue immunedysfunction syndrome (CFIDS), chronic inflammatory demyelinatingpolyneuropathy (CIDP), chronic polyarthritis, Churg-Strauss syndrome,cicatricial pemphigoid, Cogan syndrome, CREST syndrome (syndrom withCalcinosis cutis, Raynaud phenomenon, motility disorders of theesophagus, sklerodaktylia and teleangiectasia), Crohn's disease (MorbusCrohn, colitis ulcerosa), dermatitis herpetiformis during, dermatologicautoimmune diseases, dermatomyositis, Diabetes, Diabetes mellitus Type 1(type I diabetes, insuline dependent Diabetes mellitus), Diabetesmellitus Type 2 (type II diabetes), essential mixed cryoglobulinemia,essential mixed cryoglobulinemia, fibromyalgia, fibromyositis,Goodpasture syndrome (anti-GBM mediated glomerulonephritis), graftversus host disease, Guillain-Barré syndrome (GBM,Polyradikuloneuritis), haematologic autoimmune diseases, Hashimotothyroiditis, hemophilia, acquired hemophilia, hepatitis, autoimmunehepatitis, particularly autoimmune forms of chronic hepatitis,idiopathic pulmonary fibrosis (IPF), idiopathic thrombocytopenicpurpura, Immuno-thrombocytopenic purpura (Morbus Werlhof; ITP), IgAnephropathy, infertility, autoimmune infertility, juvenile rheumatoidarthritis (Morbus Still, Still syndrome), Lambert-Eaton syndrome, lichenplanus, lichen sclerosus, lupus erythematosus, systemic lupuserythematosus (SLE), lupus erythematosus (discoid form), Lyme arthritis(Lyme disease, borrelia arthritis), Ménierè's disease (Morbus Ménierè);mixed connective tissue disease (MCTD), multiple sclerosis (MS,encephalomyelitis disseminate, Charcot's disease), Myasthenia gravis(myasthenia, MG), myosits, polymyositis, neural autoimmune diseases,neurodermitis, pemphigus-vulgaris, bullous pemphigoid, scar formingpemphigoid; polyarteriitis nodosa (periarteiitis nodosa), polychondritis(panchondritis), polyglandular (autoimmune) syndrome (PGA syndrome,Schmidt's syndrome), Polymyalgia rheumatica, primary agammaglobulinemia,primary biliary cirrhosis PBC, primary autoimmune cholangitis),progressive systemic sclerosis (PSS), Psoriasis, Psoriasis vulgaris,Raynaud's phenomena, Reiter's syndrome (Morbus Reiter, urethralconjunctive synovial syndrome)), rheumatoid arthritis (RA, chronicpolyarthritis, rheumatic disease of the joints, rheumatic fever),sarcoidosis (Morbus Boeck, Besnier-Boeck-Schaumann disease), stiff-mansyndrome, Sclerodermia, Scleroderma, Sjögren's syndrome, sympatheticophtalmia; Transient gluten intolerance, transplanted organ rejection,uveitis, autoimmune uveiitis, Vasculitis, Vitiligo, (leucoderma, pieboldskin), and Wegner's disease (Morbus Wegner, Wegner's granulomatosis)

In a further preferred aspect, the inventive nucleic acid sequence asdefined herein or the inventive composition comprising a plurality ofinventive nucleic acid sequences as defined herein may be used for thepreparation of a pharmaceutical composition or a vaccine, particularlyfor purposes as defined herein.

The inventive pharmaceutical composition or vaccine may furthermore beused for the treatment of a disease or a disorder, preferably ofallergies or autoimmune diseases as defined herein.

According to a final aspect, the present invention also provides kits,particularly kits of parts. Such kits, particularly kits of parts,typically comprise as components alone or in combination with furthercomponents as defined herein at least one inventive nucleic acidsequence as defined herein, the inventive pharmaceutical composition orvaccine comprising the inventive nucleic acid sequence. The at least oneinventive nucleic acid sequence as defined herein, optionally incombination with further components as defined herein, whereby the atleast one nucleic acid of the invention is provided separately (firstpart of the kit) from at least one other part of the kit comprising oneor more other components. The inventive pharmaceutical compositionand/or the inventive vaccine may e.g. occur in one or different parts ofthe kit. As an example, e.g. at least one part of the kit may compriseat least one inventive nucleic acid sequence as defined herein, and atleast one further part of the kit at least one other component asdefined herein, e.g. at least one other part of the kit may comprise atleast one pharmaceutical composition or vaccine or a part thereof, e.g.at least one part of the kit may comprise the inventive nucleic acidsequence as defined herein, at least one further part of the kit atleast one other component as defined herein, at least one further partof the kit at least one component of the inventive pharmaceuticalcomposition or vaccine or the inventive pharmaceutical composition orvaccine as a whole, and at least one further part of the kit e.g. atleast one pharmaceutical carrier or vehicle, etc. In case the kit or kitof parts comprises a plurality of inventive nucleic acid sequences, onecomponent of the kit can comprise only one, several or all inventivenucleic acid sequences comprised in the kit. In an alternativeembodiment every/each inventive nucleic acid sequence may be comprisedin a different/separate component of the kit such that each componentforms a part of the kit. Also, more than one nucleic acid may becomprised in a first component as part of the kit, whereas one or moreother (second, third etc.) components (providing one or more other partsof the kit) may either contain one or more than one inventive nucleicacids, which may be identical or partially identical or different fromthe first component. The kit or kit of parts may furthermore containtechnical instructions with information on the administration and dosageof the inventive nucleic acid sequence, the inventive pharmaceuticalcomposition or the inventive vaccine or of any of its components orparts, e.g. if the kit is prepared as a kit of parts.

Taken together, the invention provides a nucleic acid sequencecomprising or coding for

-   -   a) a coding region, encoding at least one peptide or protein;    -   b) at least one histone stem-loop, and    -   c) a poly(A) sequence or a polyadenylation signal;    -   wherein said protein or peptide antigen is an allergenic antigen        or a fragment, variant or derivative of said allergenic antigen,        or an autoimmune self-antigen associated with an autoimmune        disease.

Preferably, the invention provides a provides a nucleic acid sequencecomprising or coding for

-   -   a) a coding region, encoding at least one peptide or protein;    -   b) at least one histone stem-loop, and    -   c) a poly(A) sequence or a polyadenylation signal;        wherein said protein or peptide antigen is an allergenic antigen        or a fragment, variant or derivative of said allergenic antigen,        or an autoimmune self-antigen asscociated with an autoimmune        disease, wherein the autoimmune disease is associated with at        least one organ system, particularly with the circulatory        system, the digestive system, the endocrine system, the        excretory system, the immune system, the integumentary system,        the muscular system, the nervous system, the reproductive        system, the respiratory system, the skeletal system, preferably        with the the cardiovascular system, the neuroendocrine system,        the musculoskeletal system or gastrointestinal system.

Preferably, the invention provides a nucleic acid sequence comprising orcoding for

-   -   a) a coding region, encoding at least one peptide or protein;    -   b) at least one histone stem-loop, and    -   c) a poly(A) sequence or a polyadenylation signal;        wherein said protein or peptide antigen is an allergenic antigen        or a fragment, variant or derivative of said allergenic antigen,        or an autoimmune self-antigen associated with an autoimmune        disease, preferably wherein said protein or peptide antigen is        an autoimmune self-antigen associated with an autoimmune disease        selected from Addison disease (autoimmune adrenalitis, Morbus        Addison), alopecia areata, Addison's anemia (Morbus Biermer),        autoimmune hemolytic anemia (AIHA), autoimmune hemolytic anemia        (AIHA) of the cold type (cold hemagglutinine disease, cold        autoimmune hemolytic anemia (AIHA) (cold agglutinin disease),        (CHAD)), autoimmune hemolytic anemia (AIHA) of the warm type        (warm AIHA, warm autoimmune haemolytic anemia (AIHA)),        autoimmune hemolytic Donath-Landsteiner anemia (paroxysmal cold        hemoglobinuria), antiphospholipid syndrome (APS),        atherosclerosis, autoimmune arthritis, arteriitis temporalis,        Takayasu arteriitis (Takayasu's disease, aortic arch disease),        temporal arteriitis/giant cell arteriitis, autoimmune chronic        gastritis, autoimmune infertility, autoimmune inner ear disease        (AIED), Basedow's disease (Morbus Basedow), Bechterew's disease        (Morbus Bechterew, ankylosing spondylitis, spondylitis        ankylosans), Behcet's syndrome (Morbus Behcet), bowel disease        including autoimmune inflammatory bowel disease (including        colitis ulcerosa (Morbus Crohn, Crohn's disease),        cardiomyopathy, particularly autoimmune cardiomyopathy,        idiopathic dilated cardiomyopathy (DCM), celiac sprue dermatitis        (gluten mediated enteropathia), chronic fatigue immune        dysfunction syndrome (CFIDS), chronic inflammatory demyelinating        polyneuropathy (CIDP), chronic polyarthritis, Churg-Strauss        syndrome, cicatricial pemphigoid, Cogan syndrome, CREST syndrome        (syndrom with Calcinosis cutis, Raynaud phenomenon, motility        disorders of the esophagus, sklerodaktylia and teleangiectasia),        Crohn's disease (Morbus Crohn, colitis ulcerosa), dermatitis        herpetiformis during, dermatologic autoimmune diseases,        dermatomyositis, Diabetes, Diabetes mellitus Type 1 (type I        diabetes, insuline dependent Diabetes mellitus), Diabetes        mellitus Type 2 (type II diabetes), essential mixed        cryoglobulinemia, essential mixed cryoglobulinemia,        fibromyalgia, fibromyositis, Goodpasture syndrome (anti-GBM        mediated glomerulonephritis), graft versus host disease,        Guillain-Barré syndrome (GBM, Polyradikuloneuritis),        haematologic autoimmune diseases, Hashimoto thyroiditis,        hemophilia, acquired hemophilia, hepatitis, autoimmune        hepatitis, particularly autoimmune forms of chronic hepatitis,        idiopathic pulmonary fibrosis (IPF), idiopathic thrombocytopenic        purpura, Immuno-thrombocytopenic purpura (Morbus Werlhof; ITP),        IgA nephropathy, infertility, autoimmune infertility, juvenile        rheumatoid arthritis (Morbus Still, Still syndrome),        Lambert-Eaton syndrome, lichen planus, lichen sclerosus, lupus        erythematosus, systemic lupus erythematosus (SLE), lupus        erythematosus (discoid form), Lyme arthritis (Lyme disease,        borrelia arthritis), Ménierè's disease (Morbus Ménierè); mixed        connective tissue disease (MCTD), multiple sclerosis (MS,        encephalomyelitis disseminate, Charcot's disease), Myasthenia        gravis (myasthenia, MG), myosits, polymyositis, neural        autoimmune diseases, neurodermitis, pemphigus vulgaris, bullous        pemphigoid, scar forming pemphigoid; polyarteriitis nodosa        (periarteiitis nodosa), polychondritis (panchondritis),        polyglandular (autoimmune) syndrome (PGA syndrome, Schmidt's        syndrome), Polymyalgia rheumatica, primary agammaglobulinemia,        primary biliary cirrhosis PBC, primary autoimmune cholangitis),        progressive systemic sclerosis (PSS), Psoriasis, Psoriasis        vulgaris, Raynaud's phenomena, Reiter's syndrome (Morbus Reiter,        urethral conjunctive synovial syndrome)), rheumatoid arthritis        (RA, chronic polyarthritis, rheumatic disease of the joints,        rheumatic fever), sarcoidosis (Morbus Boeck,        Besnier-Boeck-Schaumann disease), stiff-man syndrome,        Sclerodermia, Scleroderma, Sjögren's syndrome, sympathetic        ophtalmia; Transient gluten intolerance, transplanted organ        rejection, uveitis, autoimmune uveiitis, Vasculitis, Vitiligo,        (leucoderma, piebold skin), and Wegner's disease (Morbus Wegner,        Wegner's granulomatosis), or a fragment, variant or derivative        of said autoimmune self-antigen.

Preferably, the invention provides a nucleic acid sequence comprising orcoding for

-   -   a) a coding region, encoding at least one peptide or protein;    -   b) at least one histone stem-loop, and    -   c) a poly(A) sequence or a polyadenylation signal;        wherein said protein or peptide antigen is an allergenic antigen        or a fragment, variant or derivative of said allergenic antigen,        or an autoimmune self-antigen associated with an autoimmune        disease, preferably wherein the said protein or peptide antigen        is an autoimmune self-antigen associated with an autoimmune        disease, particularly an self-antigen selected from    -   myelin basic protein (MBP), proteolipid protein (PLP), and        myelin oligodendrocyte glycoprotein (MOG), in each case        associated with multiple sclerosis (MS);    -   CD44, preproinsulin, proinsulin, insulin, glutamic acid        decaroxylase (GAD65), tyrosine phosphatase-like insulinoma        antigen 2 (IA2), zinc transporter ((ZnT8), and heat shock        protein 60 (HSP60), in each case associated with diabetes Typ I;    -   interphotoreceptor retinoid-binding protein (IRBP) associated        with autoimmune uveitis;    -   acetylcholine receptor AchR, and insulin-like growth factor-1        receptor (IGF-1R), in each case associated with Myasthenia        gravis;    -   M-protein from beta-hemolytic streptocci (pseudo-autoantigen)        associated with Rheumatic Fever;    -   Macrophage migration inhibitory factor associated with        Arthritis;    -   Ro/La RNP complex, alpha- and beta-fodrin, islet cell        autoantigen, poly(ADP)ribose polymerase (PARP), NuMA, NOR-90,        Ro60 autoantigen, and p27 antigen, in each case associated with        Sjögren's syndrome;    -   Ro60 autoantigen, low-density lipoproteins, Sm antigens of the        U-1 small nuclear ribonucleoprotein complex (B/B′, D1, D2, D3,        E, F, G), and RNP ribonucleoproteins, in each case associated        with lupus erythematosus;    -   oxLDL, beta(2)GPI, HSP60/65, and oxLDL/beta(2)GPI, in each case        associated with Atherosclerosis;    -   cardiac beta(1)-adrenergic receptor associated with idiopathic        dilated cardiomyopathy (DCM);    -   histidyl-tRNA synthetase (HisRS) associated with myositis;    -   topoisomerase I associated with scleroderma disease.

Preferably, the invention provides a nucleic acid sequence comprising orcoding for

-   -   a) a coding region, encoding at least one peptide or protein;    -   b) at least one histone stem-loop, and    -   c) a poly(A) sequence or a polyadenylation signal;        wherein said protein or peptide antigen is an allergenic antigen        or a fragment, variant or derivative of said allergenic antigen,        or an autoimmune self-antigen asscociated with an autoimmune        disease, preferably, wherein said protein or peptide antigen is        an allergenic antigen derived from a source selected from the        list consisting of: grass pollen, tree pollen, flower pollen,        herb pollen, dust mite, mold, animals, food, and insect venom,        or a fragment, variant or derivative of said allergenic antigen.

In the present invention, if not otherwise indicated, different featuresof alternatives and embodiments may be combined with each other.Furthermore, the term “comprising” shall not be construed as meaning“consisting of”, if not specifically mentioned. However, in the contextof the present invention, term “comprising” may be substituted with theterm “consisting of”, where applicable.

FIGURES

The following Figures are intended to illustrate the invention furtherand shall not be construed to limit the present invention thereto.

FIG. 1: shows the histone stem-loop consensus sequence generated frommetazoan and protozoan stem loop sequences (as reported by Dávila López,M., & Samuelsson, T. (2008), RNA (New York, N.Y.), 14(1), 1-10.doi:10.1261/rna.782308). 4001 histone stem-loop sequences from metazoaand protozoa were aligned and the quantity of the occurring nucleotidesis indicated for every position in the stem-loop sequence. The generatedconsensus sequence representing all nucleotides present in the sequencesanalyzed is given using the single-letter nucleotide code. In additionto the consensus sequence, sequences are shown representing at least99%, 95% and 90% of the nucleotides present in the sequences analyzed.

FIG. 2: shows the histone stem-loop consensus sequence generated fromprotozoan stem loop sequences (as reported by Davila López, M., &Samuelsson, T. (2008), RNA (New York, N.Y.), 14(1), 1-10.doi:10.1261/rna.782308). 131 histone stem-loop sequences from protozoawere aligned and the quantity of the occurring nucleotides is indicatedfor every position in the stem-loop sequence. The generated consensussequence representing all nucleotides present in the sequences analyzedis given using the single-letter nucleotide code. In addition to theconsensus sequence, sequences are shown representing at least 99%, 95%and 90% of the nucleotides present in the sequences analyzed.

FIG. 3: shows the histone stem-loop consensus sequence generated frommetazoan stem loop sequences (as reported by Dávila López, M., &Samuelsson, T. (2008), RNA (New York, N.Y.), 14(1), 1-10.doi:10.1261/rna.782308). 3870 histone stem-loop sequences from metazoawere aligned and the quantity of the occurring nucleotides is indicatedfor every position in the stem-loop sequence. The generated consensussequence representing all nucleotides present in the sequences analyzedis given using the single-letter nucleotide code. In addition to theconsensus sequence, sequences are shown representing at least 99%, 95%and 90% of the nucleotides present in the sequences analyzed.

FIG. 4: shows the histone stem-loop consensus sequence generated fromvertebrate stem loop sequences (as reported by Dávila López, M., &Samuelsson, T. (2008), RNA (New York, N.Y.), 14(1), 1-10.doi:10.1261/rna.782308). 1333 histone stem-loop sequences fromvertebrates were aligned and the quantity of the occurring nucleotidesis indicated for every position in the stem-loop sequence. The generatedconsensus sequence representing all nucleotides present in the sequencesanalyzed is given using the single-letter nucleotide code. In additionto the consensus sequence, sequences are shown representing at least99%, 95% and 90% of the nucleotides present in the sequences analyzed.

FIG. 5: shows the histone stem-loop consensus sequence generated fromhuman (Homo sapiens) stem loop sequences (as reported by Dávila López,M., & Samuelsson, T. (2008), RNA (New York, N.Y.), 14(1), 1-10.doi:10.1261/rna.782308). 84 histone stem-loop sequences from humans werealigned and the quantity of the occurring nucleotides is indicated forevery position in the stem-loop sequence. The generated consensussequence representing all nucleotides present in the sequences analyzedis given using the single-letter nucleotide code. In addition to theconsensus sequence, sequences are shown representing at least 99%, 95%and 90% of the nucleotides present in the sequences analyzed.

FIGS. 6 to 19: show mRNAs from in vitro transcription.

Given are the designation and the sequence of mRNAs obtained by in vitrotranscription. The following abbreviations are used:

-   -   ppLuc (GC): GC-enriched mRNA sequence coding for Photinus        pyra/is luciferase    -   ag: 3′ untranslated region (UTR) of the alpha globin gene    -   A64: poly(A)-sequence with 64 adenylates    -   A120: poly(A)-sequence with 120 adenylates    -   histoneSL: histone stem-loop    -   aCPSL: stem loop which has been selected from a library for its        specific binding of the αCP-2KL protein    -   PolioCL: 5′ clover leaf from Polio virus genomic RNA    -   G30: poly(G) sequence with 30 guanylates    -   U30: poly(U) sequence with 30 uridylates    -   SL: unspecific/artificial stem-loop    -   N32: unspecific sequence of 32 nucleotides    -   MmMOG (wt): wild type mRNA sequence encoding murine        myelin-oligodendroglialprotein (MOG)    -   Within the sequences, the following elements are highlighted:        coding region (ORF) (capital letters), ag (bold), histoneSL        (underlined), further distinct sequences tested (italic).

FIG. 6: shows the mRNA sequence of ppLuc(GC)-ag (SEQ ID NO: 43). Bylinearization of the original vector at the restriction site immediatelyfollowing the alpha-globin 3′-UTR (ag), mRNA is obtained lacking apoly(A) sequence.

FIG. 7: shows the mRNA sequence of ppLuc(GC)-ag-A64 (SEQ ID NO: 44). Bylinearization of the original vector at the restriction site immediatelyfollowing the A64 poly(A)-sequence, mRNA is obtained ending with an A64poly(A) sequence.

FIG. 8: shows the mRNA sequence of ppLuc(GC)-ag-histoneSL (SEQ ID NO:45). The A64 poly(A) sequence was replaced by a histoneSL. The histonestem-loop sequence used in the examples was obtained from Cakmakci etal. (2008). Molecular and Cellular Biology, 28(3), 1182-1194.

FIG. 9: shows the mRNA sequence of ppLuc(GC)-ag-A64-histoneSL (SEQ IDNO: 46).

-   -   The histoneSL was appended 3′ of A64 poly(A).

FIG. 10: shows the mRNA sequence of ppLuc(GC)-ag-A120 (SEQ ID NO: 47).The A64 poly(A) sequence was replaced by an A120 poly(A) sequence.

FIG. 11: shows the mRNA sequence of ppLuc(GC)-ag-A64-ag (SEQ ID NO: 48).A second alpha-globin 3′-UTR was appended 3′ of A64 poly(A).

FIG. 12: shows the mRNA sequence of ppLuc(GC)-ag-A64-aCPSL (SEQ ID NO:49).

A stem loop was appended 3′ of A64 poly(A). The stem loop has beenselected from a library for its specific binding of the αCP-2KL protein(Thisted et at, (2001), The Journal of Biological Chemistry, 276(20),17484-17496). αCP-2KL is an isoform of αCP-2, the most stronglyexpressed αCP protein (alpha-globin mRNA poly(C) binding protein)(Makeyev et at, (2000), Genomics, 67(3), 301-316), a group of RNAbinding proteins, which bind to the alpha-globin 3′-UTR (Chkheidze etat, (1999), Molecular and Cellular Biology, 19(7), 4572-4581).

FIG. 13: shows the mRNA sequence of ppLuc(GC)-ag-A64-PolioCL (SEQ ID NO:50).

-   -   The 5′ clover leaf from Polio virus genomic RNA was appended 3′        of A64 poly(A).

FIG. 14: shows the mRNA sequence of ppLuc(GC)-ag-A64-G30 (SEQ ID NO: 51)A stretch of 30 guanylates was appended 3′ of A64 poly(A).

FIG. 15: shows the mRNA sequence of ppLuc(GC)-ag-A64-U30 (SEQ ID NO: 52)A stretch of 30 uridylates was appended 3′ of A64 poly(A).

FIG. 16: shows the mRNA sequence of ppLuc(GC)-ag-A64-SL (SEQ ID NO: 53)A stem loop was appended 3′ of A64 poly(A). The upper part of the stemand the loop were taken from (Babendure et at, (2006), RNA (New York,N.Y.), 12(5), 851-861). The stem loop consists of a 17 base pair long,CG-rich stem and a 6 base long loop.

FIG. 17: shows ppLuc(GC)-ag-A64-N32 (SEQ ID NO: 54)

-   -   By linearization of the original vector at an alternative        restriction site, mRNA is obtained with 32 additional        nucleotides following poly(A).

FIG. 18: shows the mRNA sequence of MmMOG(wt)-ag-A64-C30 (SEQ ID NO: 55)

FIG. 19: shows the mRNA sequence of MmMOG(wt)-ag-A64-C30-histoneSL (SEQID NO: 56)

FIG. 20: shows that the combination of poly(A) and histoneSL increasesprotein expression from mRNA in a synergistic manner.

-   -   The effect of poly(A) sequence, histoneSL, and the combination        of poly(A) and histoneSL on luciferase expression from mRNA was        examined. Therefore different mRNAs were electroporated into        HeLa cells. Luciferase levels were measured at 6, 24, and 48        hours after transfection. Little luciferase is expressed from        mRNA having neither poly(A) sequence nor histoneSL. Both a        poly(A) sequence or the histoneSL increase the luciferase level.        Strikingly however, the combination of poly(A) and histoneSL        further strongly increases the luciferase level, manifold above        the level observed with either of the individual elements, thus        acting synergistically. Data are graphed as mean RLU±SD        (relative light units±standard deviation) for triplicate        transfections. Specific RLU are summarized in Example 10.2.

FIG. 21: shows that the combination of poly(A) and histoneSL increasesprotein expression from mRNA irrespective of their order.

-   -   The effect of poly(A) sequence, histoneSL, the combination of        poly(A) and histoneSL, and their order on luciferase expression        from mRNA was examined. Therefore different mRNAs were        lipofected into HeLa cells.

Luciferase levels were measured at 6, 24, and 48 hours after the startof transfection. Both an A64 poly(A) sequence or the histoneSL give riseto comparable luciferase levels. Increasing the length of the poly(A)sequence from A64 to A120 or to A300 increases the luciferase levelmoderately. In contrast, the combination of poly(A) and histoneSLincreases the luciferase level much further than lengthening of thepoly(A) sequence. The combination of poly(A) and histoneSL actssynergistically as it increases the luciferase level manifold above thelevel observed with either of the individual elements. The synergisticeffect of the combination of poly(A) and histoneSL is seen irrespectiveof the order of poly(A) and histoneSL and irrespective of the length ofpoly(A) with A64-histoneSL or histoneSL-A250 mRNA. Data are graphed asmean RLU±SD for triplicate transfections. Specific RLU are summarized inExample 10.3.

FIG. 22: shows that the rise in protein expression by the combination ofpoly(A) and histoneSL is specific.

-   -   The effect of combining poly(A) and histoneSL or poly(A) and        alternative sequences on luciferase expression from mRNA was        examined. Therefore different mRNAs were electroporated into        HeLa cells. Luciferase levels were measured at 6, 24, and 48        hours after transfection. Both a poly(A) sequence or the        histoneSL give rise to comparable luciferase levels. The        combination of poly(A) and histoneSL strongly increases the        luciferase level, manifold above the level observed with either        of the individual elements, thus acting synergistically. In        contrast, combining poly(A) with any of the other sequences is        without effect on the luciferase level compared to mRNA        containing only a poly(A) sequence. Thus, the combination of        poly(A) and histoneSL acts specifically and synergistically.        Data are graphed as mean RLU±SD for triplicate transfections.        Specific RLU are summarized in Example 10.4.

FIG. 23: shows that the combination of poly(A) and histoneSL increasesprotein expression from mRNA in a synergistic manner in vivo.

-   -   The effect of poly(A) sequence, histoneSL, and the combination        of poly(A) and histoneSL on luciferase expression from mRNA in        vivo was examined. Therefore different mRNAs were injected        intradermally into mice. Mice were sacrificed 16 hours after        injection and Luciferase levels at the injection sites were        measured. Luciferase is expressed from mRNA having either a        histoneSL or a poly(A) sequence. Strikingly however, the        combination of poly(A) and histoneSL strongly increases the        luciferase level, manifold above the level observed with either        of the individual elements, thus acting synergistically. Data        are graphed as mean RLU±SEM (relative light units±standard error        of mean). Specific RLU are summarized in Example 10.5.

EXAMPLES

The following Examples are intended to illustrate the invention furtherand shall not be construed to limit the present invention thereto.

1. Generation of Histone-Stem-Loop Consensus Sequences

-   -   Prior to the experiments, histone stem-loop consensus sequences        were determined on the basis of metazoan and protozoan histone        stem-loop sequences. Sequences were taken from the supplement        provided by Lopez et al. (Dávila López, M., & Samuelsson, T.        (2008), RNA (New York, N.Y.), 14(1), 1-10.        doi:10.1261/rna.782308), who identified a large number of        natural histone stem-loop sequences by searching genomic        sequences and expressed sequence tags. First, all sequences from        metazoa and protozoa (4001 sequences), or all sequences from        protozoa (131 sequences) or alternatively from metazoa (3870        sequences), or from vertebrates (1333 sequences) or from humans        (84 sequences) were grouped and aligned. Then, the quantity of        the occurring nucleotides was determined for every position.        Based on the tables thus obtained, consensus sequences for the 5        different groups of sequences were generated representing all        nucleotides present in the sequences analyzed. In addition, more        restrictive consensus sequences were also obtained, increasingly        emphasizing conserved nucleotides

2. Preparation of DNA-Templates

-   -   A vector for in vitro transcription was constructed containing a        T7 promoter followed by a GC-enriched sequence coding for        Photinus pyra/is luciferase (ppLuc(GC)), the center part of the        3′ untranslated region (UTR) of alpha-globin (ag), and a poly(A)        sequence. The poly(A) sequence was immediately followed by a        restriction site used for linearization of the vector before in        vitro transcription in order to obtain mRNA ending in an A64        poly(A) sequence. mRNA obtained from this vector accordingly by        in vitro transcription is designated as “ppLuc(GC)-ag-A64”.    -   Linearization of this vector at alternative restriction sites        before in vitro transcription allowed to obtain mRNA either        extended by additional nucleotides 3′ of A64 or lacking A64. In        addition, the original vector was modified to include        alternative sequences. In summary, the following mRNAs were        obtained from these vectors by in vitro transcription (mRNA        sequences are given in FIGS. 6 to 17):    -   ppLuc(GC)-ag (SEQ ID NO: 43)    -   ppLuc(GC)-ag-A64 (SEQ ID NO: 44)    -   ppLuc(GC)-ag-histoneSL (SEQ ID NO: 45)    -   ppLuc(GC)-ag-A64-histoneSL (SEQ ID NO: 46)    -   ppLuc(GC)-ag-A120 (SEQ ID NO: 47)    -   ppLuc(GC)-ag-A64-ag (SEQ ID NO: 48)    -   ppLuc(GC)-ag-A64-aCPSL (SEQ ID NO: 49)    -   ppLuc(GC)-ag-A64-PolioCL (SEQ ID NO: 50)    -   ppLuc(GC)-ag-A64-G30 (SEQ ID NO: 51)    -   ppLuc(GC)-ag-A64-U30 (SEQ ID NO: 52)    -   ppLuc(GC)-ag-A64-SL (SEQ ID NO: 53)    -   ppLuc(GC)-ag-A64-N32 (SEQ ID NO: 54)    -   Furthermore DNA plasmid sequences coding for the autoimmune        self-antigen MOG were prepared accordingly as described above.    -   In summary, the following mRNAs were obtained from these vectors        by in vitro transcription (mRNA sequences are given in FIGS. 18        to 19):    -   MmMOG(wt)-ag-A64-C30 (SEQ ID NO: 55)    -   MmMOG(wt)-ag-A64-C30-histoneSL (SEQ ID NO: 56)

3. In Vitro Transcription

The DNA-template according to Example 2 was linearized and transcribedin vitro using T7-Polymerase. The DNA-template was then digested byDNase-treatment. All mRNA-transcripts contained a 5′-CAP structureobtained by adding an excess ofN7-Methyl-Guanosine-5′-Triphosphate-5′-Guanosine to the transcriptionreaction. mRNA thus obtained was purified and resuspended in water.

4. Enzymatic Adenylation of mRNA

-   -   Two mRNAs were enzymatically adenylated:    -   ppLuc(GC)-ag-A64 and ppLuc(GC)-ag-histoneSL.    -   To this end, RNA was incubated with E. coli Poly(A)-polymerase        and ATP (Poly(A) Polymerase Tailing Kit, Epicentre, Madison,        USA) following the manufacturer's instructions. mRNA with        extended poly(A) sequence was purified and resuspended in water.        The length of the poly(A) sequence was determined via agarose        gel electrophoresis. Starting mRNAs were extended by        approximately 250 adenylates, the mRNAs obtained are designated        as    -   ppLuc(GC)-ag-A300 and ppLuc(GC)-ag-histoneSL -A250,        respectively.        5. Luciferase Expression by mRNA Electroporation    -   HeLa cells were trypsinized and washed in opti-MEM. 1×10⁵ cells        in 200 μl of opti-MEM each were electroporated with 0.5 μg of        ppLuc-encoding mRNA. As a control, mRNA not coding for ppLuc was        electroporated separately. Electroporated cells were seeded in        24-well plates in 1 ml of RPMI 1640 medium. 6, 24, or 48 hours        after transfection, medium was aspirated and cells were lysed in        200 μpl of lysis buffer (25 mM Tris, pH 7.5 (HCl), 2 mM EDTA,        10% glycerol, 1% Triton X-100, 2 mM DTT, 1 mM PMSF). Lysates        were stored at −20° C. until ppLuc activity was measured.        6. Luciferase Expression by mRNA Lipofection    -   HeLa cells were seeded in 96 well plates at a density of 2×10⁴        cells per well. The following day, cells were washed in opti-MEM        and then transfected with 0.25 μg of Lipofectin-complexed        ppLuc-encoding mRNA in 150 μl of opti-MEM. As a control, mRNA        not coding for ppLuc was lipofected separately. In some wells,        opti-MEM was aspirated and cells were lysed in 200 μl of lysis        buffer 6 hours after the start of transfection. In the remaining        wells, opti-MEM was exchanged for RPMI 1640 medium at that time.        In these wells, medium was aspirated and cells were lysed in 200        μl of lysis buffer 24 or 48 hours after the start of        transfection. Lysates were stored at −20° C. until ppLuc        activity was measured.

7. Luciferase Measurement

-   -   ppLuc activity was measured as relative light units (RLU) in a        BioTek SynergyHT plate reader at 5 seconds measuring time using        50 μl of lysate and 200 μl of luciferin buffer (25 mM        Glycylglycin, pH 7.8 (NaOH), 15 mM MgSO₄, 2 mM ATP, 75 μM        luciferin). Specific RLU were calculated by subtracting RLU of        the control RNA from total RLU.        8. Luciferase Expression by Intradermal mRNA Injection        (Luciferase Expression In Vivo)    -   Mice were anaesthetized with a mixture of Rompun and Ketavet.        Each ppLuc-encoding mRNA was injected intradermally (0.5 μg of        mRNA in 50 μl per injection). As a control, mRNA not coding for        ppLuc was injected separately. 16 hours after injection, mice        were sacrificed and tissue collected. Tissue samples were flash        frozen in liquid nitrogen and lysed in a tissue lyser (Qiagen)        in 800 μl of lysis buffer (25 mM Tris, pH 7.5 (HCl), 2 mM EDTA,        10% glycerol, 1% Triton X-100, 2 mM DTT, 1 mM PMSF).        Subsequently samples were centrifuged at 13500 rpm at 4° C. for        10 minutes. Lysates were stored at −80° C. until ppLuc activity        was measured (see 7. luciferase measurement).

9. Experimental Autoimmune Encephalomyelitis (EAE):

-   -   Among autoimmune diseases a particular prominent example is        multiple sclerosis (MS). MS is one of the most frequently        occurring neurological diseases in young adults and affects        about 1 million people worldwide. It is commonly accepted that        MS is triggered by an autoimmune reaction against myelin        components (e.g. myelin basic protein (MBP), proteolipidprotein        (PLP), myelin-oligodendroglialprotein (MOG), etc.), of the human        body, wherein a combination of genetic predisposition and        environmental factors is assumed to be involved in the onset of        MS. A frequently investigated model for multiple sclerosis is        the mouse model of experimental autoimmune encephalomyelitis        (EAE).    -   EAE is triggered in C57BL/6 mice at day 0 by administration of        recombinant MOG protein and an adjuvant cocktail. The cocktail        contains recombinant MOG protein and CFA, the administration is        carried out via injection with complete Freund's adjuvans. A        sensibilization is achieved by administration of Pertussis        toxin. The mice are dazed with KetaminNetranquil prior to        administration.    -   In order to weaken the symptoms of disease, mice are vaccinated        with the mRNAs according to SEQ ID NOs: 55 and 56 after        triggering EAE in the mouse model. Therefore, mRNAs encoding MOG        (the construct accoding to SEQ ID NOs: 55 and 56) are        administered via injection in two cycles with one week        difference between both cycles. In each of those cycles four        injections are carried out (on days 1, 2, 3, 4, 15, 16, 17 and        18), i.e., in each cycle administration is carried out at four        subsequent days per week. The mRNAs according to SEQ ID NOs: 55        and 56 are formulated with protamine and administered in a        concentration of 10 μg per ear per injection (intradermally).    -   The mice and the progression of the disease is monitored over        the whole time period in regular intervals. The progression of        the disease is determined upon following EAE score, which uses        the extent of paralysis as a measure for progression of disease:

0 no symptoms; 0.5 end of tail paresis; 1 complete tail paresis; 2 weakdistal paresis; 2.5 body reflexes restricted upon paresis; 3 mediumparesis, abdominal muscles involved; 3.5 paresis of hind legs, bellytouches the ground; 4 paraplegia or tetraparesis; 4.5 tetraplegia ortetraparesis + incontinence; 5 moribund.

10 Results 10.1 Histone Stem-Loop Sequences:

-   -   In order to characterize histone stem-loop sequences, sequences        from metazoa and protozoa (4001 sequences), or from protozoa        (131 sequences) or alternatively from metazoa (3870 sequences),        or from vertebrates (1333 sequences) or from humans (84        sequences) were grouped and aligned. Then, the quantity of the        occurring nucleotides was determined for every position. Based        on the tables thus obtained, consensus sequences for the 5        different groups of sequences were generated representing all        nucleotides present in the sequences analyzed. Within the        consensus sequence of metazoa and protozoa combined, 3        nucleotides are conserved, a T/U in the loop and a G and a C in        the stem, forming a base pair. Structurally, typically a 6        base-pair stem and a loop of 4 nucleotides is formed. However,        deviating structures are common: Of 84 human histone stem-loops,        two contain a stem of only 5 nucleotides comprising 4 base-pairs        and one mismatch. Another human histone stem-loop contains a        stem of only 5 base-pairs. Four more human histone stem-loops        contain a 6 nucleotide long stem, but include one mismatch at        three different positions, respectively. Furthermore, four human        histone stem-loops contain one wobble base-pair at two different        positions, respectively. Concerning the loop, a length of 4        nucleotides seems not to be strictly required, as a loop of 5        nucleotides has been identified in D. discoideum.    -   In addition to the consensus sequences representing all        nucleotides present in the sequences analyzed, more restrictive        consensus sequences were also obtained, increasingly emphasizing        conserved nucleotides. In summary, the following sequences were        obtained:    -   (Cons): represents all nucleotides present    -   (99%): represents at least 99% of all nucleotides present    -   (95%): represents at least 95% of all nucleotides present    -   (90%): represents at least 90% of all nucleotides present    -   The results of the analysis of histone stem-loop sequences are        summarized in the following Tables 1 to 5 (see also FIGS. 1 to        5):

TABLE 1Metazoan and protozoan histone stem-loop consensus sequence: (based onan alignment of 4001 metazoan and protozoan histone stem-loop sequences) (see also FIG. 1) < < < < < < • • • • > # A 2224 1586 3075 2872 1284 184    0   13   12    9    1   47   59    0  675 3818 # T  172  188  47  205   19    6    0  569 1620  199 3947 3830 3704 4001  182    1 #C 1557 2211  875  918 2675  270    0 3394 2342 3783   51  119  227    03140    7 # G   25   16    4    6   23 3541 4001   25   27   10    2   5   11    0    4  175 Cons N* N* N N N N G N N N N N N T N N 99% H*H* H H V V G Y Y Y Y H H T H R 95% M* H* M H M S G Y Y Y T T Y T M A 90%M* M* M M M S G Y Y C T T T T M A > > > > > # A 195 1596 523 0 14 372761 771 2012 2499 # T 21 15 11 0 179 8 64 557 201 690 # C 50 31 16 40013543 154 3870 2636 1744 674 # G 3735 2359 3451 0 265 112 4 37 43 138Cons N N N C N N N N* N* N* 99% V V R C B V H H* N* N* 95% R R R C S M CH* H* H* 90% G R R C S A C H* M* H*

TABLE 2 Protozoan histone stem-loop consensus sequence: (based onan alignment of 131 protozoan histone stem-loop sequences)(see also FIG. 2) < < < < < < • • • • > > > > > > # A 52 32 71 82 76  13  0 12 12  9  1 46  3   0 75 82 53 79 20   0   4 94 17 35 74 56 # T 2032 37 21  8   3   0 21 85 58 86 70 65 131 28  1 17 13 10   0  15  7 3132 20 28 # C 45 59 20 25 38   0   0 86  8 54 42 13 58   0 27  2  6 31 10131 112  5 82 58 30 40 # G 14  8  3  3  9 115 131 12 26 10  2  2  5   0 1 46 55  8 91   0   0 25  1  6  7  7 Cons N* N* N N N D G N N N N N N TN N N N N C H N N N* N* N* 99% N* N* N N N D G N N N B N N T H V N N N CH N H N* N* N* 95% N* N* H H N R G N N N Y H B T H R D N N C Y D H H* N*N* 90% N* H* H H V R G N D B Y H Y T H R D H N C Y R H H* H* H*

TABLE 3 Metazoan histone stem-loop consensus sequence: (based onan alignment of 3870 (including 1333 vertebrate sequences)metazoan histone stem-loop sequences) (see also FIG. 3) < < < < < < • •• • > # A 2172 1554 3004 2790 1208  171    0    1    0    0    0    1  56    0  600 3736 # T  152  156   10  184   11    3    0  548 1535 141 3861 3760 3639 3870  154    0 # C 1512 2152  855  893 2637  270   0 3308 2334 3729    9  106  169    0 3113    5 # G   11    8    1   3   14 3426 3870   13    1    0    0    3    6    0    3  129 Cons N*N* N N N N G N B Y Y N N T N    V 99% H* H* M H M V G Y Y Y T Y H T H   R 95% M* M* M M M S G V V C T T Y T M    A 90% M* M* M M M S G Y Y CT T T T M    A > > > > > # A  142 1517  503    0   10 3633   44  7361938 2443 # T    4    2    1    0  164    1   33  525  181  662 # C   44   0    6 3870 3431  149 3788 2578 1714  634 # G 3680 2351 3360    0 265   87    3   31   36  131 Cons N D N C N N N N* N* N* 99% V R R C BV M H* H* N* 95% G R R C S M C H* H* H* 90% G R R C S A C H* M* H*

TABLE 4 Vertebrate histone stem-loop consensus sequence: (based onan alignment of 1333 vertebrate histone stem-loop sequences)(see also FIG. 4) < < < < < < • • • • > # A 661  146 1315 1323 920    8   0    1    0    0    0    1    4    0  441 1333 # T  63  121    2    2  6    2    0   39 1217    2 1331 1329 1207 1333   30    0 # C 601 1062  16    6 403    1    0 1293  116 1331    2    0  121    0  862    0 # G  8    4    0    2   4 1322 1333    0    0    0    0    3    1    0    0   0 Cons N* N* H N N N G H Y Y V D N T H A 99% H* H* M A M G G V Y C TT V T H A 95% H* H* A A M G G C V C T T V T M A 90% M* M* A A M G G C TC T T T T M A > > > > > # A    0 1199   21    0    1 1126   26   81  380 960 # T    1    0    1    0    2    1   22   91   91    12 # C    2   0    0 1333 1328  128 1284 1143  834  361 # G 1330  134 1311    0   2   78    1   18   28    0 Cons B R D C N N N N* N* H* 99% G R R C CV H N* N* M* 95% G R G C C V C W H* M* 90% G R G C C M C Y* M* M*

TABLE 5 Homo sapiens histone stem-loop consensus sequence: (based onan alignment of 84 human histone stem-loop sequences) (see also FIG. 5)< < < < < < • • • • > > > > > > # A 10 17 84 84 76  1  0  1  0  0  0  1 0  0 12 84  0 65  3  0  0 69  5  0 10 64 # T  8  6  0  0  2  2  0  1 67 0 84 80 81 84  5  0  0  0  0  0  0  0  4 25 24  3 # C 62 61  0  0  6  0 0 82 17 84  0  0  3  0 67  0  1  0  0 84 84  5 75 57 44 17 # G  4  0  0 0  0 81 84  0  0  0  0  3  0  0  0  0 83 19 81  0  0 10  0  2  6  0Cons N* H* A A H D C H Y C T D Y T H A S R R C C V H B* N* H* 99% N* H*A A H D C H Y C T D Y T H A S R R C C V H B* N* H* 95% H* H* A A M G G CY C T T T T H A G R G C C V M Y* N* M* 90% H* M* A A A G C C Y C T T T TM A C R C C C R M Y* H* M*

-   -   Wherein the used abbreviations were defined as followed:

abbreviation Nucleotide bases remark G G Guanine A A Adenine T T ThymineU U Uracile C C Cytosine R G or A Purine Y T/U or C Pyrimidine M A or CAmino K G or T/U Keto S G or C Strong (3H bonds) W A or T/U Weak (2Hbonds) H A or C or T/U Not G B G or T/U or C Not A V G or C or A Not T/UD G or A or T/U Not C N G or C or T/U or A Any base * present or notBase may be present or not10.2 the Combination of Poly(A) and histoneSL Increases ProteinExpression from mRNA in a Synergistic Manner.

-   -   To investigate the effect of the combination of poly(A) and        histoneSL on protein expression from mRNA, mRNAs with different        sequences 3′ of the alpha-globin 3′-UTR were synthesized: mRNAs        either ended just 3′ of the 3′-UTR, thus lacking both poly(A)        sequence and histoneSL, or contained either an A64 poly(A)        sequence or a histoneSL instead, or both A64 poly(A) and        histoneSL 3′ of the 3′-UTR. Luciferase-encoding mRNAs or control        mRNA were electroporated into HeLa cells. Luciferase levels were        measured at 6, 24, and 48 hours after transfection (see        following Table 6 and FIG. 20).

TABLE 6 RLU at RLU at RLU at mRNA 6 hours 24 hours 48 hoursppLuc(GC)-ag-A64-histoneSL 466553 375169 70735 ppLuc(GC)-ag-histoneSL50947 3022 84 ppLuc(GC)-ag-A64 10471 19529 4364 ppLuc(GC)-ag 997 217 42

-   -   Little luciferase was expressed from mRNA having neither poly(A)        sequence nor histoneSL. Both a poly(A) sequence or the histoneSL        increased the luciferase level to a similar extent. Either mRNA        gave rise to a luciferase level much higher than did mRNA        lacking both poly(A) and histoneSL. Strikingly however, the        combination of poly(A) and histoneSL further strongly increased        the luciferase level, manifold above the level observed with        either of the individual elements. The magnitude of the rise in        luciferase level due to combining poly(A) and histoneSL in the        same mRNA demonstrates that they are acting synergistically.    -   The synergy between poly(A) and histoneSL was quantified by        dividing the signal from poly(A)-histoneSL mRNA (+/+) by the sum        of the signals from histoneSL mRNA (−/+) plus poly(A) mRNA (+/−)        (see following Table 7).

TABLE 7 RLU at RLU at RLU at A64 histoneSL 6 hours 24 hours 48 hours + +466553 375169 70735 − + 50947 3022 84 + − 10471 19529 4364 Synergy 7.616.6 15.9

-   -   The factor thus calculated specifies how much higher the        luciferase level from mRNA combining poly(A) and histoneSL is        than would be expected if the effects of poly(A) and histoneSL        were purely additive. The luciferase level from mRNA combining        poly(A) and histoneSL was up to 16.6 times higher than if their        effects were purely additive. This result confirms that the        combination of poly(A) and histoneSL effects a markedly        synergistic increase in protein expression.        10.3 the Combination of Poly(A) and histoneSL Increases Protein        Expression from mRNA Irrespective of their Order.    -   The effect of the combination of poly(A) and histoneSL might        depend on the length of the poly(A) sequence and the order of        poly(A) and histoneSL. Thus, mRNAs with increasing poly(A)        sequence length and mRNA with poly(A) and histoneSL in reversed        order were synthesized: Two mRNAs contained 3′ of the 3′-UTR        either an A120 or an A300 poly(A) sequence. One further mRNA        contained 3′ of the 3′-UTR first a histoneSL followed by an A250        poly(A) sequence. Luciferase-encoding mRNAs or control mRNA were        lipofected into HeLa cells. Luciferase levels were measured at        6, 24, and 48 hours after the start of transfection (see        following Table 8 and FIG. 21).

TABLE 8 RLU at RLU at RLU at mRNA 6 hours 24 hours 48 hoursppLuc(GC)-ag-histoneSL-A250 98472 734222 146479ppLuc(GC)-ag-A64-histoneSL 123674 317343 89579 ppLuc(GC)-ag-histoneSL7291 4565 916 ppLuc(GC)-ag-A300 4357 38560 11829 ppLuc(GC)-ag-A120 437145929 10142 ppLuc(GC)-ag-A64 1928 26781 537

-   -   Both an A64 poly(A) sequence or the histoneSL gave rise to        comparable luciferase levels. In agreement with the previous        experiment did the combination of A64 and histoneSL strongly        increase the luciferase level, manifold above the level observed        with either of the individual elements. The magnitude of the        rise in luciferase level due to combining poly(A) and histoneSL        in the same mRNA demonstrates that they are acting        synergistically. The synergy between A64 and histoneSL was        quantified as before based on the luciferase levels of        A64-histoneSL, A64, and histoneSL mRNA (see following Table 9).        The luciferase level from mRNA combining A64 and histoneSL was        up to 61.7 times higher than if the effects of poly(A) and        histoneSL were purely additive.

TABLE 9 RLU at RLU at RLU at A64 histoneSL 6 hours 24 hours 48 hours + +123674 317343 89579 − + 7291 4565 916 + − 1928 26781 537 Synergy 13.410.1 61.7

-   -   In contrast, increasing the length of the poly(A) sequence from        A64 to A120 or to A300 increased the luciferase level only        moderately (see Table 8 and FIG. 19). mRNA with the longest        poly(A) sequence, A300, was also compared to mRNA in which a        poly(A) sequence of similar length was combined with the        histoneSL, histoneSL-A250. In addition to having a long poly(A)        sequence, the order of histoneSL and poly(A) is reversed in this        mRNA relative to A64-histoneSL mRNA. The combination of A250 and        histoneSL strongly increased the luciferase level, manifold        above the level observed with either histoneSL or A300. Again,        the synergy between A250 and histoneSL was quantified as before        comparing RLU from histoneSL-A250 mRNA to RLU from A300 mRNA        plus histoneSL mRNA (see following Table 10). The luciferase        level from mRNA combining A250 and histoneSL was up to 17.0        times higher than if the effects of poly(A) and histoneSL were        purely additive.

TABLE 10 RLU at RLU at RLU at histoneSL A250/A300 6 hours 24 hours 48hours + + 98472 734222 146479 + − 7291 4565 916 − + 4357 38560 11829Synergy 8.5 17.0 11.5

-   -   In summary, a highly synergistic effect of the combination of        histoneSL and poly(A) on protein expression from mRNA has been        demonstrated for substantially different lengths of poly(A) and        irrespective of the order of poly(A) and histoneSL.        10.4 the Rise in Protein Expression by the Combination of        Poly(A) and histoneSL is specific    -   To investigate whether the effect of the combination of poly(A)        and histoneSL on protein expression from mRNA is specific, mRNAs        with alternative sequences in combination with poly(A) were        synthesized: These mRNAs contained 3′ of A64 one of seven        distinct sequences, respectively. Luciferase-encoding mRNAs or        control mRNA were electroporated into HeLa cells. Luciferase        levels were measured at 6, 24, and 48 hours after transfection        (see following Table 11 and FIG. 22).

TABLE 11 RLU at RLU at RLU at mRNA 6 hours 24 hours 48 hoursppLuc(GC)-ag-A64-N32 33501 38979 2641 ppLuc(GC)-ag-A64-SL 28176 20364874 ppLuc(GC)-ag-A64-U30 41632 54676 3408 ppLuc(GC)-ag-A64-G30 4676349210 3382 ppLuc(GC)-ag-A64-PolioCL 46428 26090 1655ppLuc(GC)-ag-A64-aCPSL 34176 53090 3338 ppLuc(GC)-ag-A64-ag 18534 18194989 ppLuc(GC)-ag-A64-histoneSL 282677 437543 69292ppLuc(GC)-ag-histoneSL 27597 3171 0 ppLuc(GC)-ag-A64 14339 48414 9357

-   -   Both a poly(A) sequence or the histoneSL gave rise to comparable        luciferase levels. Again, the combination of poly(A) and        histoneSL strongly increased the luciferase level, manifold        above the level observed with either of the individual elements,        thus acting synergistically. In contrast, combining poly(A) with        any of the alternative sequences was without effect on the        luciferase level compared to mRNA containing only a poly(A)        sequence. Thus, the combination of poly(A) and histoneSL        increases protein expression from mRNA in a synergistic manner,        and this effect is specific.        10.5 the Combination of Poly(A) and histoneSL Increases Protein        Expression from mRNA in a Synergistic Manner In Vivo.    -   To investigate the effect of the combination of poly(A) and        histoneSL on protein expression from mRNA in vivo,        Luciferase-encoding mRNAs with different sequences 3′ of the        alpha-globin 3′-UTR or control mRNA were injected intradermally        into mice: mRNAs contained either an A64 poly(A) sequence or a        histoneSL instead, or both A64 poly(A) and histoneSL 3′ of the        3′-UTR. Luciferase levels were measured at 16 hours after        injection (see following Table 12 and FIG. 23).

TABLE 12 RLU at mRNA 16 hours ppLuc(GC)-ag-A64-histoneSL 38081ppLuc(GC)-ag-histoneSL 137 ppLuc(GC)-ag-A64 4607

-   -   Luciferase was expressed from mRNA having either a histoneSL or        a poly(A) sequence. Strikingly however, the combination of        poly(A) and histoneSL further strongly increased the luciferase        level, manifold above the level observed with either of the        individual elements. The magnitude of the rise in luciferase        level due to combining poly(A) and histoneSL in the same mRNA        demonstrates that they are acting synergistically.    -   The synergy between poly(A) and histoneSL was quantified by        dividing the signal from poly(A)-histoneSL mRNA (+/+) by the sum        of the signals from histoneSL mRNA (−/+) plus poly(A) mRNA (+/−)        (see following Table 13).

TABLE 13 RLU at A64 histoneSL 16 hours + + 38081 − + 137 + − 4607Synergy 8.0

-   -   The factor thus calculated specifies how much higher the        luciferase level from mRNA combining poly(A) and histoneSL is        than would be expected if the effects of poly(A) and histoneSL        were purely additive. The luciferase level from mRNA combining        poly(A) and histoneSL was 8 times higher than if their effects        were purely additive. This result confirms that the combination        of poly(A) and histoneSL effects a markedly synergistic increase        in protein expression in vivo.

1-22. (canceled)
 23. A nucleic acid molecule comprising: a) apolypeptide coding region, encoding an autoimmune disease antigen or anallergenic antigen; b) at least one histone stem-loop, and c) a poly(A)sequence or a polyadenylation signal.
 24. The nucleic acid moleculeaccording to claim 23, wherein the polypeptide coding region encodes anautoimmune disease antigen.
 25. The nucleic acid molecule according toclaim 24, wherein the autoimmune disease is associated the circulatorysystem, the digestive system, the endocrine system, the excretorysystem, the immune system, the integumentary system, the muscularsystem, the nervous system, the reproductive system, the respiratorysystem, the skeletal system, preferably with the cardiovascular system,the neuroendocrine system, the musculoskeletal system or thegastrointestinal system.
 26. The nucleic acid molecule of claim 24,wherein the autoimmune disease antigen is associated with an autoimmunedisease selected from the group consisting of Addison disease, alopeciaareata, Addison's anemia, autoimmune hemolytic anemia (AIHA), autoimmunehemolytic anemia (AIHA), cold hemagglutinine disease, cold autoimmunehemolytic anemia (CHAD), warm AIHA, autoimmune hemolyticDonath-Landsteiner anemia, antiphospholipid syndrome (APS),atherosclerosis, autoimmune arthritis, arteriitis temporalis, Takayasuarteriitis, temporal arteriitis/giant cell arteriitis, autoimmunechronic gastritis, autoimmune infertility, autoimmune inner ear disease(AIED), Basedow's disease, Bechterew's disease, Behcet's syndrome,autoimmune inflammatory bowel disease, Crohn's disease, autoimmunecardiomyopathy, idiopathic dilated cardiomyopathy (DCM), celiac spruedermatitis, chronic fatigue immune dysfunction syndrome (CFIDS), chronicinflammatory demyelinating polyneuropathy (CIDP), chronic polyarthritis,Churg-Strauss syndrome, cicatricial pemphigoid, Cogan syndrome, CRESTsyndrome, dermatitis herpetiformis, dermatologic autoimmune disease,dermatomyositis, Diabetes mellitus Type 1, Diabetes mellitus Type 2,essential mixed cryoglobulinemia, fibromyalgia, fibromyositis,Goodpasture syndrome, graft versus host disease, Guillain-Barrésyndrome, haematologic autoimmune diseases, Hashimoto thyroiditis,hemophilia, acquired hemophilia, hepatitis, autoimmune hepatitis,autoimmune chronic hepatitis, idiopathic pulmonary fibrosis (IPF),idiopathic thrombocytopenic purpura, Immuno-thrombocytopenic purpura(ITP), IgA nephropathy, infertility, autoimmune infertility, juvenilerheumatoid arthritis, Lambert-Eaton syndrome, lichen planus, lichensclerosus, lupus erythematosus, systemic lupus erythematosus (SLE),lupus erythematosus (discoid form), Lyme arthritis, Ménierè's disease,mixed connective tissue disease (MCTD), multiple sclerosis (MS),encephalomyelitis disseminate, Charcot's disease, Myasthenia gravis(MG), myosits, polymyositis, neural autoimmune diseases, neurodermitis,pemphigus vulgaris, bullous pemphigoid, scar forming pemphigoid,polyarteriitis nodosa (periarteiitis nodosa), polychondritis(panchondritis), polyglandular (autoimmune) syndrome (PGA syndrome),Polymyalgia rheumatica, primary agammaglobulinemia, primary biliarycirrhosis PBC, primary autoimmune cholangitis, progressive systemicsclerosis (PSS), Psoriasis, Psoriasis vulgaris, Raynaud's phenomena,Reiter's syndrome, rheumatoid arthritis (RA), chronic polyarthritis,rheumatic disease of the joints, rheumatic fever, sarcoidosis, stiff-mansyndrome, Sclerodermia, Scleroderma, Sjögren's syndrome, sympatheticophtalmia, Transient gluten intolerance, transplanted organ rejection,uveitis, autoimmune uveiitis, Vasculitis, Vitiligo, leucoderma, pieboldskin, and Wegner's disease.
 27. The nucleic acid molecule of claim 24,wherein the autoimmune disease antigen is an autoimmune self-antigenselected from the group consisting of myelin basic protein (MBP),proteolipid protein (PLP), myelin oligodendrocyte glycoprotein (MOG),CD44, preproinsulin, proinsulin, insulin, glutamic acid decaroxylase(GAD65), tyrosine phosphatase-like insulinoma antigen 2 (IA2), zinctransporter (ZnT8), heat shock protein 60 (HSP60), interphotoreceptorretinoid-binding protein (IRBP), acetylcholine receptor AchR,insulin-like growth factor-1 receptor (IGF-1R), m-protein frombeta-hemolytic streptocci (pseudo-autoantigen), macrophage migrationinhibitory factor, Ro/La RNP complex, alpha-fodrin, islet cellautoantigen, beta-fodrin, islet cell autoantigen, poly(ADP)ribosepolymerase (PARP), NuMA, NOR-90, Ro60 autoantigen, p27 antigen, Ro60autoantigen, a low-density lipoprotein, a Sm antigens of the U-1 smallnuclear ribonucleoprotein complex, a RNP ribonucleoproteins, oxLDL,beta(2)GPI, HSP60/65, oxLDL/beta(2)GPI, cardiac beta(1)-adrenergicreceptor, histidyl-tRNA synthetase (HisRS), and topoisomerase I.
 28. Thenucleic acid molecule according to claim 23, wherein the polypeptidecoding region encodes an allergenic antigen.
 29. The nucleic acidmolecule of claim 28, wherein the allergenic antigen is derived from asource selected from the list consisting of grass pollen, tree pollen,flower pollen, herb pollen, dust mite, mold, animals, food, and insectvenom.
 30. The nucleic acid molecule of claim 23, wherein the moleculedoes not comprise a sequence encoding a reporter protein, a marker orselection protein.
 31. The nucleic acid molecule of claim 23, whereinthe molecule does not comprise sequence encoding a reporter protein, amarker or selection protein.
 32. The nucleic acid molecule of claim 23,wherein the nucleic acid is a RNA.
 33. The nucleic acid molecule ofclaim 23, wherein the poly(A) sequence comprises a sequence of about 25to about 400 adenosine nucleotides.
 34. The nucleic acid molecule ofclaim 23, wherein the polyadenylation signal comprises the consensussequence NN(U/T)ANA.
 35. The nucleic acid molecule of claim 23, whereinat least one guanosine, uridine, adenosine, thymidine, or cytidineposition of the nucleic acid molecule is substituted with an analogue ofthese nucleotides selected from2-amino-6-chloropurineriboside-5′-triphosphate,2-aminoadenosine-5′-triphosphate, 2-thiocytidine-5′-triphosphate,2-thiouridine-5′-triphosphate, 4-thiouridine-5′-triphosphate,5-aminoallylcytidine-5′-triphosphate,5-aminoallyluridine-5′-triphosphate, 5-bromocytidine-5′-triphosphate,5-bromouridine-5′-triphosphate, 5-iodocytidine-5′-triphosphate,5-iodouridine-5′-triphosphate, 5-methylcytidine-5′-triphosphate,5-methyluridine-5′-triphosphate, 6-azacytidine-5′-triphosphate,6-azauridine-5′-triphosphate, 6-chloropurineriboside-5′-triphosphate,7-deazaadenosine-5′-triphosphate, 7-deazaguanosine-5′-triphosphate,8-azaadenosine-5′-triphosphate, 8-azidoadenosine-5′-triphosphate,benzimidazole-riboside-5′-triphosphate,N1-methyladenosine-5′-triphosphate, N1-methylguanosine-5′-triphosphate,N6-methyladenosine-5′-triphosphate, 06-methylguanosine-5′-triphosphate,pseudouridine-5′-triphosphate, or puromycin-5′-triphosphate, andxanthosine-5′-triphosphate.
 36. The nucleic acid sequence molecule ofclaim 23, wherein the G/C content of the polypeptide coding region isincreased compared with the G/C content of the coding region of awild-type nucleic acid encoding the autoimmune disease antigen or theallergenic antigen.
 37. The nucleic acid molecule of claim 32, whereinthe RNA comprises a 5′ cap structure and a poly(A) sequence of about 25to about 400 adenosine nucleotides.
 38. The nucleic acid molecule ofclaim 23, wherein the nucleic acid molecule comprises a sequence of atleast 10 consecutive cytidines.
 39. The nucleic acid molecule of claim23, wherein the nucleic acid molecule further comprises a stabilizingsequence from the alpha globin 3′ UTR, positioned 3′ relative to thepolypeptide coding region of the nucleic acid molecule.
 40. Apharmaceutical composition comprising a nucleic acid molecule of claim23 and a pharmaceutically acceptable carrier.
 41. The pharmaceuticalcomposition of claim 40, further comprising an adjuvant.
 42. Thepharmaceutical composition of claim 40, wherein the composition furthercomprises a cationic or polycationic compound in complex with thenucleic acid molecule.
 43. The pharmaceutical composition of claim 42,wherein the polycationic polypeptide in complex with the nucleic acidmolecule comprise protamine.
 44. A method for expressing a polypeptidein a patient comprising administering an effective amount of a nucleicacid molecule to a patient, the nucleic acid molecule comprising: a) apolypeptide coding region, encoding an autoimmune disease antigen or anallergenic antigen; b) at least one histone stem-loop, and c) a poly(A)sequence or a polyadenylation signal, thereby expressing the autoimmunedisease antigen or the allergenic antigen in the patient.